Fused-tricyclic inhibitors of KRAS and methods of use thereof

ABSTRACT

Compounds having activity as inhibitors of G12C mutant KRAS protein are provided. The compounds have the following structure (I):(I) or a pharmaceutically acceptable salt, stereoisomer or prodrug thereof, wherein A is a heterocyclic or heteroaryl ring, and R 1 , R 2a , R 2b , R 2c , R 3a , R 3b , R 4a , R 4b , A, G 1 , G 2 , L 1 , L 2 , m 1 , m 2 , and E are as defined herein. Methods associated with preparation and use of such compounds, pharmaceutical compositions comprising such compounds and methods to modulate the activity of G12C mutant KRAS protein for treatment of disorders, such as cancer, are also provided.

BACKGROUND Technical Field

The present invention is generally directed to novel compounds andmethods for their preparation and use as therapeutic or prophylacticagents, for example for treatment of cancer.

Description of the Related Art

RAS represents a group of closely related monomeric globular proteins of189 amino acids (21 kDa molecular mass) which are associated with theplasma membrane and which bind either GDP or GTP. RAS acts as amolecular switch. When RAS contains bound GDP, it is in the resting oroff position and is “inactive”. In response to exposure of the cell tocertain growth promoting stimuli, RAS is induced to exchange its boundGDP for a GTP. With GTP bound, RAS is “switched on” and is able tointeract with and activate other proteins (its “downstream targets”).The RAS protein itself has a very low intrinsic ability to hydrolyze GTPback to GDP, thus turning itself into the off state. Switching RAS offrequires extrinsic proteins termed GTPase-activating proteins (GAPs)that interact with RAS and greatly accelerate the conversion of GTP toGDP. Any mutation in RAS which affects its ability to interact with GAPor to convert GTP back to GDP will result in a prolonged activation ofthe protein and consequently a prolonged signal to the cell telling itto continue to grow and divide. Because these signals result in cellgrowth and division, overactive RAS signaling may ultimately lead tocancer.

Structurally, RAS proteins contain a G domain which is responsible forthe enzymatic activity of RAS—the guanine nucleotide binding and thehydrolysis (GTPase reaction). It also contains a C-terminal extension,known as the CAAX box, which may be post-translationally modified and isresponsible for targeting the protein to the membrane. The G domain isapproximately 21-25 kDa in size and it contains a phosphate binding loop(P-loop). The P-loop represents the pocket where the nucleotides arebound in the protein, and this is the rigid part of the domain withconserved amino acid residues which are essential for nucleotide bindingand hydrolysis (Glycine 12, Threonine 26 and Lysine 16). The G domainalso contains the so called Switch I (residues 30-40) and Switch II(residues 60-76) regions, both of which are the dynamic parts of theprotein which are often represented as the “spring-loaded” mechanismbecause of their ability to switch between the resting and loaded state.The key interaction is the hydrogen bonds formed by Threonine-35 andglycine-60 with the γ-phosphate of GTP which maintain Switch 1 andSwitch 2 regions respectively in their active conformation. Afterhydrolysis of GTP and release of phosphate, these two relax into theinactive GDP conformation.

The most notable members of the RAS subfamily are HRAS, KRAS and NRAS,mainly for being implicated in many types of cancer. However, there aremany other members including DIRAS1; DIRAS2; DIRAS3; ERAS; GEM; MRAS;NKIRAS1; NKIRAS2; NRAS; RALA; RALB; RAP1A; RAP1B; RAP2A; RAP2B; RAP2C;RASD1; RASD2; RASL10A; RASL10B; RASL11A; RASL11B; RASL12; REM1; REM2;RERG; RERGL; RRAD; RRAS and RRAS2.

Mutations in any one of the three main isoforms of RAS (HRAS, NRAS, orKRAS) genes are among the most common events in human tumorigenesis.About 30% of all human tumors are found to carry some mutation in RASgenes. Remarkably, KRAS mutations are detected in 25-30% of tumors. Bycomparison, the rates of oncogenic mutation occurring in the NRAS andHRAS family members are much lower (8% and 3% respectively). The mostcommon KRAS mutations are found at residue G12 and G13 in the P-loop andat residue Q61.

G12C is a frequent mutation of KRAS gene (glycine-12 to cysteine). Thismutation had been found in about 13% of cancer occurrences, about 43% oflung cancer occurrences, and in almost 100% of MYH-associates polyposis(familial colon cancer syndrome). However targeting this gene with smallmolecules is a challenge.

Accordingly, while progress has been made in this field, there remains aneed in the art for improved compounds and methods for treatment ofcancer, for example by inhibition of KRAS, HRAS or NRAS. The presentinvention fulfills this need and provides further related advantages.

BRIEF SUMMARY

In brief, the present invention provides compounds, includingstereoisomers, pharmaceutically acceptable salts, tautomers and prodrugsthereof, which are capable of modulating G12C mutant KRAS, HRAS and/orNRAS proteins. In some instances, the compounds act as electrophileswhich are capable of forming a covalent bond with the cysteine residueat position 12 of a KRAS, HRAS or NRAS G12C mutant protein. Methods foruse of such compounds for treatment of various diseases or conditions,such as cancer, are also provided.

In one embodiment, compounds having the following structure (I) areprovided:

or a pharmaceutically acceptable salt, stereoisomer or prodrug thereof,wherein R¹, R^(2a), R^(2b), R^(2c), R^(3a), R^(3b), R^(4a), R^(4b), A,G¹, G², L¹, L², m¹, m² and E are as defined herein. Pharmaceuticalcompositions comprising one or more compounds of structure (I) and apharmaceutically acceptable carrier are also provided in various otherembodiments.

In other embodiments, the present invention provides a method fortreatment of cancer, the method comprising administering an effectiveamount of a pharmaceutical composition comprising any one or more of thecompounds of structure (I) to a subject in need thereof.

Other provided methods include a method for regulating activity of aKRAS, HRAS or NRAS G12C mutant protein, the method comprising reactingthe KRAS, HRAS or NRAS G12C mutant protein with any one of the compoundsof structure (I). In other embodiments, a method for inhibitingproliferation of a cell population, the method comprising contacting thecell population with any one of the compounds of structure (I) is alsoprovided.

In other embodiments, the invention is directed to a method for treatinga disorder mediated by a KRAS, HRAS or NRAS G12C mutation in a subjectin need thereof, the method comprising:

determining if the subject has a KRAS, HRAS or NRAS G12C mutation; and

if the subject is determined to have the KRAS, HRAS or NRAS G12Cmutation, then administering to the subject a therapeutically effectiveamount of a pharmaceutical composition comprising any one or morecompounds of structure (I).

In still more embodiments, the invention is directed to a method forpreparing a labeled KRAS, HRAS or NRAS G12C mutant protein, the methodcomprising reacting the KRAS, HRAS or NRAS G12C mutant with a compoundof structure (I), to result in the labeled KRAS, HRAS or NRAS G12Cprotein.

These and other aspects of the invention will be apparent upon referenceto the following detailed description.

BRIEF DESCRIPTION OF THE DRAWINGS

In the figures, identical reference numbers identify similar elements.The sizes and relative positions of elements in the figures are notnecessarily drawn to scale and some of these elements are arbitrarilyenlarged and positioned to improve figure legibility. Further, theparticular shapes of the elements as drawn are not intended to conveyany information regarding the actual shape of the particular elements,and have been solely selected for ease of recognition in the figures.

FIG. 1 illustrates the enzymatic activity of RAS.

FIG. 2 depicts a signal transduction pathway for RAS.

FIG. 3 shows some common oncogenes, their respective tumor type andcumulative mutation frequencies (all tumors).

DETAILED DESCRIPTION

In the following description, certain specific details are set forth inorder to provide a thorough understanding of various embodiments of theinvention. However, one skilled in the art will understand that theinvention may be practiced without these details.

Unless the context requires otherwise, throughout the presentspecification and claims, the word “comprise” and variations thereof,such as, “comprises” and “comprising” are to be construed in an open,inclusive sense, that is as “including, but not limited to”.

Reference throughout this specification to “one embodiment” or “anembodiment” means that a particular feature, structure or characteristicdescribed in connection with the embodiment is included in at least oneembodiment of the present invention. Thus, the appearances of thephrases “in one embodiment” or “in an embodiment” in various placesthroughout this specification are not necessarily all referring to thesame embodiment. Furthermore, the particular features, structures, orcharacteristics may be combined in any suitable manner in one or moreembodiments.

Unless defined otherwise, all technical and scientific terms used hereinhave the same meaning as is commonly understood by one of skill in theart to which this invention belongs. As used in the specification andclaims, the singular form “a”, “an” and “the” include plural referencesunless the context clearly dictates otherwise.

“Amidinyl” refers to a radical of the form —(C═NR_(a))NR_(b)R_(c),wherein R_(a), R_(b) and R_(c) are each independently H or C₁-C₆ alkyl.

“Amino” refers to the —NH₂ radical.

“Aminylsulfone” refers to the —S(O)₂NH₂ radical.

“Carboxy” or “carboxyl” refers to the —CO₂H radical.

“Cyano” refers to the —CN radical.

“Guanidinyl” refers to a radical of the form—NR_(d)(C═NR_(a))NR_(b)R_(c), wherein R_(a), R_(b), R_(c), and R_(d) areeach independently H or C₁-C₆ alkyl.

“Hydroxy” or “hydroxyl” refers to the —OH radical.

“Imino” refers to the ═NH substituent.

“Nitro” refers to the —NO₂ radical.

“Oxo” refers to the ═O substituent.

“Thioxo” refers to the ═S substituent.

“Alkyl” refers to a straight or branched hydrocarbon chain radicalconsisting solely of carbon and hydrogen atoms, which is saturated orunsaturated (i.e., contains one or more double and/or triple bonds),having from one to twelve carbon atoms (C₁-C₁₂ alkyl), preferably one toeight carbon atoms (C₁-C₈ alkyl) or one to six carbon atoms (C₁-C₆alkyl), and which is attached to the rest of the molecule by a singlebond, e.g., methyl, ethyl, n-propyl, 1-methylethyl (iso-propyl),n-butyl, n-pentyl, 1,1-dimethylethyl (t-butyl), 3-methylhexyl,2-methylhexyl, ethenyl, prop-1-enyl, but-1-enyl, pent-1-enyl,penta-1,4-dienyl, ethynyl, propynyl, butynyl, pentynyl, hexynyl, and thelike. Alkyl includes alkenyls (one or more carbon-carbon double bonds)and alkynyls (one or more carbon-carbon triple bonds such as ethynyl andthe like). “Amidinylalkyl” refers to an alkyl group comprising at leastone amidinyl substituent. “Guanidinylalkyl” refers to an alkyl groupcomprising at least one guanidinyl substituent. Unless stated otherwisespecifically in the specification, an alkyl, amidinylalkyl and/orguanidinylalkyl group is optionally substituted.

“Alkylene” or “alkylene chain” refers to a straight or branched divalenthydrocarbon chain linking the rest of the molecule to a radical group,consisting solely of carbon and hydrogen, which is saturated orunsaturated (i.e., contains one or more double and/or triple bonds), andhaving from one to twelve carbon atoms, e.g., methylene, ethylene,propylene, n-butylene, ethenylene, propenylene, n-butenylene,propynylene, n-butynylene, and the like. The alkylene chain is attachedto the rest of the molecule through a single or double bond and to theradical group through a single or double bond. The points of attachmentof the alkylene chain to the rest of the molecule and to the radicalgroup can be through one carbon or any two carbons within the chain.Unless stated otherwise specifically in the specification, an alkylenechain is optionally substituted.

“Alkylcycloalkyl” refers to a radical of the formula —R_(b)R_(d) whereR_(b) is cycloalkyl as defined herein and R_(d) is an alkyl radical asdefined above. Unless stated otherwise specifically in thespecification, a alkylcycloalkyl group is optionally substituted.

“Alkoxy” refers to a radical of the formula —OR_(a) where R_(a) is analkyl radical as defined above containing one to twelve carbon atoms.“Amidinylalkyloxy” refers to an alkoxy group comprising at least oneamidinyl substituent on the alkyl group. “Guanidinylalkyloxy” refers toan alkoxy group comprising at least one guanidinyl substituent on thealkyl group. “Alkylcarbonylaminylalkyloxy” refers to an alkoxy groupcomprising at least one alkylcarbonylaminyl substituent on the alkylgroup. “Heterocyclylalkyloxy” refers to an alkoxy group comprising atleast one heterocyclyl substituent on the alkyl group.“Heteroarylalkyloxy” refers to an alkoxy group comprising at least oneheteroaryl substituent on the alkyl group. “Aminylalkyloxy” refers to analkoxy group comprising at least one substituent of the form—NR_(a)R_(b), where R_(a) and R_(b) are each independently H or C₁-C₆alkyl, on the alkyl group. Unless stated otherwise specifically in thespecification, an alkoxy, amidinylalkyloxy, guanidinylalkyloxy,alkylcarbonylaminylalkyloxy, heterocyclylalkyloxy, heteroarlyalkyloxyand/or aminylalkyloxy group is optionally substituted.

“Alkoxyalkyl” refers to a radical of the formula —R_(b)OR_(a) whereR_(a) is an alkyl radical as defined above containing one to twelvecarbon atoms and R_(b) is an alkylene radical as defined abovecontaining one to twelve carbon atoms. Unless stated otherwisespecifically in the specification, an alkoxyalkyl group is optionallysubstituted.

“Alkoxycarbonyl” refers to a radical of the formula —C(═O)OR_(a) whereR_(a) is an alkyl radical as defined above containing one to twelvecarbon atoms. Unless stated otherwise specifically in the specification,an alkoxycarbonyl group is optionally substituted.

“Aryloxy” refers to a radical of the formula —OR_(a) where R_(a) is anaryl radical as defined herein. Unless stated otherwise specifically inthe specification, an aryloxy group is optionally substituted.

“Alkylaminyl” refers to a radical of the formula —NHR_(a) or—NR_(a)R_(a) where each R_(a) is, independently, an alkyl radical asdefined above containing one to twelve carbon atoms. A “haloalkylaminyl”group is an alkylaminyl group comprising at least one halo substituenton the alkyl group. A “hydroxylalkylaminyl” group is an alkylaminylgroup comprising at least one hydroxyl substituent on the alkyl group. A“amindinylalkylaminyl” group is an alkylaminyl group comprising at leastone amidinyl substituent on the alkyl group. A “guanidinylalkylaminyl”group is an alkylaminyl group comprising at least one guanidinylsubstituent on the alkyl group. Unless stated otherwise specifically inthe specification, an alkylaminyl, haloalkylaminyl, hydroxylalkylaminyl,amidinylalkylaminyl and/or guanidinylalkylaminyl group is optionallysubstituted.

“Aminylalkyl” refers to an alkyl group comprising at least one aminylsubstituent (—NR_(a)R_(b) wherein R_(a) and R_(b) are each independentlyH or C₁-C₆ alkyl). The aminyl substituent can be on a tertiary,secondary or primary carbon. Unless stated otherwise specifically in thespecification, an aminylalkyl group is optionally substituted.

“Aminylalkylaminyl” refers to a radical of the formula —NR_(a)R_(b)wherein R_(a) is H or C₁-C₆ alkyl and R_(b) is aminylalkyl. Unlessstated otherwise specifically in the specification, an aminylalkylaminylgroup is optionally substituted.

“Aminylalkoxy” refers to a radical of the formula —OR_(a)NH₂ whereinR_(a) is alkylene. Unless stated otherwise specifically in thespecification, an aminylalkoxy group is optionally substituted.

“Alkylaminylalkoxy” refers to a radical of the formula—OR_(a)NR_(b)R_(c) wherein R_(a) is alkylene and R_(b) and R_(c) areeach independently H or C₁-C₆ alkyl, provided one of R_(b) or R_(c) isC₁-C₆ alkyl. Unless stated otherwise specifically in the specification,an alkylaminylalkoxy group is optionally substituted.

“Alkylcarbonylaminyl” refers to a radical of the formula —NH(C═O)R_(a)where R_(a) is an alkyl radical as defined above containing one totwelve carbon atoms. Unless stated otherwise specifically in thespecification, an alkylcarbonylaminyl group is optionally substituted.An alkenylcarbonylaminyl is an alkylcarbonylaminyl containing at leastone carbon-carbon double bond. An alkenylcarbonylaminyl group isoptionally substituted.

“Alkylcarbonylaminylalkoxy” refers to a radical of the formula—OR_(b)NH(C═O)R_(a) where R_(a) is an alkyl radical as defined abovecontaining one to twelve carbon atoms and R_(b) is alkylene. Unlessstated otherwise specifically in the specification, analkylcarbonylaminylalkoxy group is optionally substituted.

“Alkylaminylalkyl” refers to an alkyl group comprising at least onealkylaminyl substituent. The alkylaminyl substituent can be on atertiary, secondary or primary carbon. Unless stated otherwisespecifically in the specification, an alkylaminylalkyl group isoptionally substituted.

“Aminylcarbonyl” refers to a radical of the formula —C(═O)NR_(a)R_(b)where R_(a) and R_(b) are each independently H or alkyl. Unless statedotherwise specifically in the specification, an aminylcarbonyl group isoptionally substituted.

“Alkylaminylcarbonyl” refers to a radical of the formula—C(═O)NR_(a)R_(b), where R_(a) and R_(b) are each independently H oralkyl, provided at least one of R_(a) or R_(b) is alkyl. Unless statedotherwise specifically in the specification, an alkylaminylcarbonylgroup is optionally substituted.

“Aminylcarbonylalkyl” refers to a radical of the formula—R_(c)C(═O)NR_(a)R_(b), where R_(a) and R_(b) are each independently Hor alkyl and R_(c) is alkylene. Unless stated otherwise specifically inthe specification, an aminylcarbonylalkyl group is optionallysubstituted.

“Aminylcarbonycycloalkyl” refers to a radical of the formula—R_(c)C(═O)NR_(a)R_(b), where R_(a) and R_(b) are each independently Hor alkyl and R_(c) is cycloalkyl. Unless stated otherwise specificallyin the specification, an aminylcarbonylcycloalkyl group is optionallysubstituted.

“Aryl” refers to a carbocyclic ring system radical comprising hydrogen,6 to 18 carbon atoms and at least one aromatic ring. For purposes ofthis invention, the aryl radical is a monocyclic, bicyclic, tricyclic ortetracyclic ring system, which may include fused or bridged ringsystems. Aryl radicals include, but are not limited to, aryl radicalsderived from aceanthrylene, acenaphthylene, acephenanthrylene,anthracene, azulene, benzene, chrysene, fluoranthene, fluorene,as-indacene, s-indacene, indane, indene, naphthalene, phenalene,phenanthrene, pleiadene, pyrene, and triphenylene. Unless statedotherwise specifically in the specification, the term “aryl” or theprefix “ar-” (such as in “aralkyl”) is meant to include aryl radicalsthat are optionally substituted.

“Aralkyl” refers to a radical of the formula —R_(b)—R_(c) where R_(b) isan alkylene chain as defined above and R_(c) is one or more arylradicals as defined above, for example, benzyl, diphenylmethyl and thelike. Unless stated otherwise specifically in the specification, anaralkyl group is optionally substituted.

“Arylalkyloxy” refers to a radical of the formula —OR_(b)—R_(c) whereR_(b) is an alkylene chain as defined above and R_(c) is one or morearyl radicals as defined above, for example, benzyl, diphenylmethyl andthe like. Unless stated otherwise specifically in the specification, anaryllkyloxy group is optionally substituted.

“Arylalkylaminyl” refers to a radical of the formula—N(R_(a))R_(b)—R_(c) where R_(a) is H or C₁-C₆ alkyl, R_(b) is analkylene chain as defined above and R_(c) is one or more aryl radicalsas defined above, for example, benzyl, diphenylmethyl and the like.Unless stated otherwise specifically in the specification, anarylalkylaminyl group is optionally substituted.

“Carboxyalkyl” refers to a radical of the formula —R_(b)—R_(c) whereR_(b) is an alkylene chain as defined above and R_(c) is a carboxy groupas defined above. Unless stated otherwise specifically in thespecification, carboxyalkyl group is optionally substituted.

“Cyanoalkyl” refers to a radical of the formula —R_(b)—R_(c) where R_(b)is an alkylene chain as defined above and R_(c) is a cyano group asdefined above. Unless stated otherwise specifically in thespecification, a cyanoalkyl group is optionally substituted.

“Carbocyclic” or “carbocycle” refers to a ring system, wherein each ofthe ring atoms are carbon.

“Cycloalkyl” refers to a stable non-aromatic monocyclic or polycycliccarbocyclic radical consisting solely of carbon and hydrogen atoms,which may include fused or bridged ring systems, having from three tofifteen carbon atoms, preferably having from three to ten carbon atoms,and which is saturated or unsaturated and attached to the rest of themolecule by a single bond. Monocyclic radicals include, for example,cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, andcyclooctyl. Polycyclic radicals include, for example, adamantyl,norbornyl, decalinyl, 7,7-dimethyl-bicyclo[2.2.1]heptanyl, and the like.A “cycloalkenyl” is a cycloalkyl comprising one or more carbon-carbondouble bonds within the ring. Unless otherwise stated specifically inthe specification, a cycloalkyl (or cycloalkenyl) group is optionallysubstituted.

“Cyanocycloalkyl” refers to a radical of the formula —R_(b)—R_(c) whereR_(b) is cycloalkyl and R_(c) is a cyano group as defined above. Unlessstated otherwise specifically in the specification, a cyanocycloalkylgroup is optionally substituted.

“Cycloalkylaminylcarbonyl” refers to a radical of the formula—C(═O)NR_(a)R_(b), where R_(a) and R_(b) are each independently H orcycloalkyl, provided at least one of R_(a) or R_(b) is cycloalkyl.Unless stated otherwise specifically in the specification, ncycloalkylaminylcarbonyl group is optionally substituted.

“Cycloalkylalkyl” refers to a radical of the formula —R_(b)R_(d) whereR_(b) is an alkylene chain as defined above and R_(d) is a cycloalkylradical as defined above. Unless stated otherwise specifically in thespecification, a cycloalkylalkyl group is optionally substituted.

“Fused” refers to any ring structure described herein which is fused toan existing ring structure in the compounds of the invention. When thefused ring is a heterocyclyl ring or a heteroaryl ring, any carbon atomon the existing ring structure which becomes part of the fusedheterocyclyl ring or the fused heteroaryl ring is replaced with anitrogen atom.

“Halo” or “halogen” refers to bromo, chloro, fluoro or iodo.

“Haloalkyl” refers to an alkyl radical, as defined above, that issubstituted by one or more halo radicals, as defined above, e.g.,trifluoromethyl, difluoromethyl, trichloromethyl, 2,2,2-trifluoroethyl,1,2-difluoroethyl, 3-bromo-2-fluoropropyl, 1,2-dibromoethyl, and thelike. Unless stated otherwise specifically in the specification, ahaloalkyl group is optionally substituted.

“Halolkoxy” refers to a radical of the formula —OR_(a) where R_(a) is ahaloalkyl radical as defined herein containing one to twelve carbonatoms. Unless stated otherwise specifically in the specification, ahaloalkoxy group is optionally substituted.

“Heterocyclyl” or “heterocyclic ring” refers to a stable 3- to18-membered non-aromatic ring radical having one to twelve carbon atoms(e.g., two to twelve) and from one to six heteroatoms selected from thegroup consisting of nitrogen, oxygen and sulfur. Unless stated otherwisespecifically in the specification, the heterocyclyl radical is amonocyclic, bicyclic, tricyclic or tetracyclic ring system, which mayinclude fused, spirocyclic (“spiro-heterocyclyl”) and/or bridged ringsystems; and the nitrogen, carbon or sulfur atoms in the heterocyclylradical is optionally oxidized; the nitrogen atom is optionallyquaternized; and the heterocyclyl radical is partially or fullysaturated. Examples of such heterocyclyl radicals include, but are notlimited to, dioxolanyl, thienyl[1,3]dithianyl, decahydroisoquinolyl,imidazolinyl, imidazolidinyl, isothiazolidinyl, isoxazolidinyl,morpholinyl, octahydroindolyl, octahydroisoindolyl, 2-oxopiperazinyl,2-oxopiperidinyl, 2-oxopyrrolidinyl, oxazolidinyl, piperidinyl,piperazinyl, 4-piperidonyl, pyrrolidinyl, pyrazolidinyl, quinuclidinyl,thiazolidinyl, tetrahydrofuryl, trithianyl, tetrahydropyranyl,thiomorpholinyl, thiamorpholinyl, 1-oxo-thiomorpholinyl, and1,1-dioxo-thiomorpholinyl. Unless stated otherwise specifically in thespecification. “Heterocyclyloxy” refers to a heterocyclyl group bound tothe remainder of the molecule via an oxygen bond (—O—).“Heterocyclylaminyl” refers to a heterocyclyl group bound to theremainder of the molecule via a nitrogen bond (—NR_(a)—, where R_(a) isH or C₁-C₆ alkyl). Unless stated otherwise specifically in thespecification, a heterocyclyl, heterocyclyloxy and/or heterocyclylaminylgroup is optionally substituted.

“N-heterocyclyl” refers to a heterocyclyl radical as defined abovecontaining at least one nitrogen and where the point of attachment ofthe heterocyclyl radical to the rest of the molecule is through anitrogen atom in the heterocyclyl radical. Unless stated otherwisespecifically in the specification, a N-heterocyclyl group is optionallysubstituted.

“Heterocyclylalkyl” refers to a radical of the formula —R_(b)R_(e) whereR_(b) is an alkylene chain as defined above and R_(e) is a heterocyclylradical as defined above, and if the heterocyclyl is anitrogen-containing heterocyclyl, the heterocyclyl is optionallyattached to the alkyl radical at the nitrogen atom. Unless statedotherwise specifically in the specification, a heterocyclylalkyl groupis optionally substituted.

“Heterocyclylalkyloxy” refers to a radical of the formula —OR_(b)R_(e)where R_(b) is an alkylene chain as defined above and R_(e) is aheterocyclyl radical as defined above, and if the heterocyclyl is anitrogen-containing heterocyclyl, the heterocyclyl is optionallyattached to the alkyl radical at the nitrogen atom. Unless statedotherwise specifically in the specification, a heterocyclylalkyloxygroup is optionally substituted.

“Heterocyclylalkylaminyl” refers to a radical of the formula—N(R_(c))R_(b)R_(e) where R_(b) is an alkylene chain as defined aboveand R_(e) is a heterocyclyl radical as defined above, and if theheterocyclyl is a nitrogen-containing heterocyclyl, the heterocyclyl isoptionally attached to the alkyl radical at the nitrogen atom, R_(c) isH or C₁-C₆ alkyl. Unless stated otherwise specifically in thespecification, a heterocyclylalkylaminyl group is optionallysubstituted.

“Heteroaryl” refers to a 5- to 14-membered ring system radicalcomprising hydrogen atoms, one to thirteen ring carbon atoms, one to sixring heteroatoms selected from the group consisting of nitrogen, oxygenand sulfur, and at least one aromatic ring. For purposes of thisinvention, the heteroaryl radical may be a monocyclic, bicyclic,tricyclic or tetracyclic ring system, which may include fused or bridgedring systems; and the nitrogen, carbon or sulfur atoms in the heteroarylradical may be optionally oxidized; the nitrogen atom may be optionallyquaternized. Examples include, but are not limited to, azepinyl,acridinyl, benzimidazolyl, benzothiazolyl, benzindolyl, benzodioxolyl,benzofuranyl, benzooxazolyl, benzothiazolyl, benzothiadiazolyl,benzo[b][1,4]dioxepinyl, 1,4-benzodioxanyl, benzonaphthofuranyl,benzoxazolyl, benzodioxolyl, benzodioxinyl, benzopyranyl,benzopyranonyl, benzofuranyl, benzofuranonyl, benzothienyl(benzothiophenyl), benzotriazolyl, benzo[4,6]imidazo[1,2-a]pyridinyl,carbazolyl, cinnolinyl, dibenzofuranyl, dibenzothiophenyl, furanyl,furanonyl, isothiazolyl, imidazolyl, indazolyl, indolyl, indazolyl,isoindolyl, indolinyl, isoindolinyl, isoquinolyl, indolizinyl,isoxazolyl, naphthyridinyl, oxadiazolyl, 2-oxoazepinyl, oxazolyl,oxiranyl, 1-oxidopyridinyl, 1-oxidopyrimidinyl, 1-oxidopyrazinyl,1-oxidopyridazinyl, 1-phenyl-1H-pyrrolyl, phenazinyl, phenothiazinyl,phenoxazinyl, phthalazinyl, pteridinyl, purinyl, pyrrolyl, pyrazolyl,pyridinyl, pyrazinyl, pyrimidinyl, pyridazinyl, quinazolinyl,quinoxalinyl, quinolinyl, quinuclidinyl, isoquinolinyl,tetrahydroquinolinyl, thiazolyl, thiadiazolyl, triazolyl, tetrazolyl,triazinyl, and thiophenyl (i.e. thienyl). “Heteroaryloxy” refers to aheteroaryl group bound to the remainder of the molecule via an oxygenbond (—O—). “Heteroarylaminyl” refers to a heteroaryl group bound to theremainder of the molecule via a nitrogen bond (—NR_(a)—, where R_(a) isH or C₁-C₆ alkyl). Unless stated otherwise specifically in thespecification, a heteroaryl, heteroaryloxy and/or heteroarylaminyl groupis optionally substituted.

“N-heteroaryl” refers to a heteroaryl radical as defined abovecontaining at least one nitrogen and where the point of attachment ofthe heteroaryl radical to the rest of the molecule is through a nitrogenatom in the heteroaryl radical. Unless stated otherwise specifically inthe specification, an N-heteroaryl group is optionally substituted.

“Heteroarylalkyl” refers to a radical of the formula —R_(b)R_(f) whereR_(b) is an alkylene chain as defined above and R_(f) is a heteroarylradical as defined above. Unless stated otherwise specifically in thespecification, a heteroarylalkyl group is optionally substituted.

“Heteroarylalkyloxy” refers to a radical of the formula —OR_(b)R_(f)where R_(b) is an alkylene chain as defined above and R_(f) is aheteroaryl radical as defined above, and if the heteroaryl is anitrogen-containing heterocyclyl, the heterocyclyl is optionallyattached to the alkyl radical at the nitrogen atom. Unless statedotherwise specifically in the specification, a heteroarylalkyloxy groupis optionally substituted.

“Heteroarylalkylaminyl” refers to a radical of the formula—NR_(c)R_(b)R_(f) where R_(b) is an alkylene chain as defined above andR_(f) is a heteroaryl radical as defined above, and if the heteroaryl isa nitrogen-containing heterocyclyl, the heterocyclyl is optionallyattached to the alkyl radical at the nitrogen atom, and R_(c) is H orC₁-C₆ alkyl. Unless stated otherwise specifically in the specification,a heteroarylalkylaminyl group is optionally substituted. “Hydroxylalkyl”refers to an alkyl group comprising at least one hydroxyl substituent.The —OH substituent may be on a primary, secondary or tertiary carbon.Unless stated otherwise specifically in the specification, ahydroxylalkyl group is optionally substituted. “Hydroxylalkylaminyl” isan alkylaminyl groups comprising at least one —OH substituent, which ison a primary, secondary or tertiary carbon. Unless stated otherwisespecifically in the specification, a hydroxylalkylaminyl group isoptionally substituted.

“Phosphate” refers to the —OP(═O)(R_(a))R_(b) group, where R_(a) is OH,O⁻ or OR_(c) and R_(b) is OH, O⁻, OR_(c), or a further phosphate group(e.g., to form a di- or triphosphate), wherein R_(c) is a counter ion(e.g., Na+ and the like).

“Phosphoalkoxy” refers to an alkoxy group, as defined herein, which issubstituted with at least one phosphate group, as defined herein. Unlessstated otherwise specifically in the specification, an phosphoalkoxygroup is optionally substituted.

“Thioalkyl” refers to a radical of the formula —SR_(a) where R_(a) is analkyl radical as defined above containing one to twelve carbon atoms.Unless stated otherwise specifically in the specification, a thioalkylgroup is optionally substituted.

The term “substituted” used herein means any of the above groups (e.g.,alkyl, alkylene, alkylcycloalkyl, alkoxy, amidinylalkyloxy,guanidinylalkyloxy, alkylcarbonylaminylalkyloxy, heterocyclylalkyloxy,heteroarylalkyloxy, aminylalkyloxy, alkoxyalkyl, alkoxycarbonyl,haloalkylaminyl, hydroxylalkylaminyl, amidinylalkylaminyl,guanidinylalkylaminyl, aminylalkyl, aminylalkylaminyl, aminylalkoxy,alkylaminylalkoxy aryloxy, alkylaminyl, alkylcarbonylaminyl,alkylaminylalkyl, aminylcarbonyl, alkylaminylcarbonyl,alkylcarbonylaminylalkoxy, aminylcarbonylalkyl,aminylcarbonycycloalkylalkyl, thioalkyl, aryl, aralkyl, arylalkyloxy,arylalkylaminyl, carboxyalkyl, cyanoalkyl, cycloalkyl, cycloalkyloxy,cycloalkylaminyl, cyanocycloalkyl, cycloalkylaminylcarbonyl,cycloalkylalkyl, haloalkyl, haloalkoxy, heterocyclyl, heterocyclyloxy,heterocyclylaminyl, N-heterocyclyl, heterocyclylalkyl,heterocyclylalkyloxy, heterocyclylalkylaminyl, heteroaryl, N-heteroaryl,heteroarylalkyl, heteroarylalkyloxy, heteroarylalkylaminyl,hydroxylalkylaminyl, phosphoalkoxy and/or hydroxylalkyl) wherein atleast one hydrogen atom (e.g., 1, 2, 3 or all hydrogen atoms) isreplaced by a bond to a non-hydrogen atom such as, but not limited to: ahalogen atom such as F, Cl, Br, and I; an oxygen atom in groups such ashydroxyl groups, alkoxy groups, and ester groups; a sulfur atom ingroups such as thiol groups, thioalkyl groups, sulfone groups, sulfonylgroups, and sulfoxide groups; a nitrogen atom in groups such as amines,amides, alkylamines, dialkylamines, arylamines, alkylarylamines,diarylamines, N-oxides, imides, and enamines; a silicon atom in groupssuch as trialkylsilyl groups, dialkylarylsilyl groups, alkyldiarylsilylgroups, and triarylsilyl groups; and other heteroatoms in various othergroups. “Substituted” also means any of the above groups in which one ormore hydrogen atoms are replaced by a higher-order bond (e.g., a double-or triple-bond) to a heteroatom such as oxygen in oxo, carbonyl,carboxyl, and ester groups; and nitrogen in groups such as imines,oximes, hydrazones, and nitriles. For example, “substituted” includesany of the above groups in which one or more hydrogen atoms are replacedwith —NR_(g)R_(h), —NR_(g)C(═O)R_(h), —NR_(g)C(═O)NR_(g)R_(h),—NR_(g)C(═O)OR_(h), —NR_(g)SO₂R_(h), —OC(═O)NR_(g)R_(h), —OR_(g),—SR_(g), —SOR_(g), —SO₂R_(g), —OSO₂R_(g), —SO₂OR_(g), ═NSO₂R_(g), and—SO₂NR_(g)R_(h). “Substituted also means any of the above groups inwhich one or more hydrogen atoms are replaced with —C(═O)R_(g),—C(═O)OR_(g), —C(═O)NR_(g)R_(h), —CH₂SO₂R_(g), —CH₂SO₂NR_(g)R_(h). Inthe foregoing, R_(g) and R_(h) are the same or different andindependently hydrogen, alkyl, alkoxy, alkylaminyl, thioalkyl, aryl,aralkyl, cycloalkyl, cycloalkylalkyl, haloalkyl, heterocyclyl,N-heterocyclyl, heterocyclylalkyl, heteroaryl, N-heteroaryl and/orheteroarylalkyl. “Substituted” further means any of the above groups inwhich one or more hydrogen atoms are replaced by a bond to an aminyl,cyano, hydroxyl, imino, nitro, oxo, thioxo, halo, alkyl, alkoxy,alkylaminyl, thioalkyl, aryl, aralkyl, cycloalkyl, cycloalkylalkyl,haloalkyl, heterocyclyl, N-heterocyclyl, heterocyclylalkyl, heteroaryl,N-heteroaryl and/or heteroarylalkyl group. In addition, each of theforegoing substituents may also be optionally substituted with one ormore of the above substituents.

“Electrophile” or “electrophilic moiety” is any moiety capable ofreacting with a nucleophile (e.g., a moiety having a lone pair ofelectrons, a negative charge, a partial negative charge and/or an excessof electrons, for example a —SH group). Electrophiles typically areelectron poor or comprise atoms which are electron poor. In certainembodiments an electrophile contains a positive charge or partialpositive charge, has a resonance structure which contains a positivecharge or partial positive charge or is a moiety in which delocalizationor polarization of electrons results in one or more atom which containsa positive charge or partial positive charge. In some embodiments, theelectrophiles comprise conjugated double bonds, for example anα,β-unsaturated carbonyl or α,β-unsaturated thiocarbonyl compound.

The term “effective amount” or “therapeutically effective amount” refersto that amount of a compound described herein that is sufficient toeffect the intended application including but not limited to diseasetreatment, as defined below. The therapeutically effective amount mayvary depending upon the intended treatment application (in vivo), or thesubject and disease condition being treated, e.g., the weight and age ofthe subject, the severity of the disease condition, the manner ofadministration and the like, which can readily be determined by one ofordinary skill in the art. The term also applies to a dose that willinduce a particular response in target cells, e.g. reduction of plateletadhesion and/or cell migration. The specific dose will vary depending onthe particular compounds chosen, the dosing regimen to be followed,whether it is administered in combination with other compounds, timingof administration, the tissue to which it is administered, and thephysical delivery system in which it is carried.

As used herein, “treatment” or “treating” refer to an approach forobtaining beneficial or desired results with respect to a disease,disorder or medical condition including but not limited to a therapeuticbenefit and/or a prophylactic benefit. By therapeutic benefit is meanteradication or amelioration of the underlying disorder being treated.Also, a therapeutic benefit is achieved with the eradication oramelioration of one or more of the physiological symptoms associatedwith the underlying disorder such that an improvement is observed in thesubject, notwithstanding that the subject may still be afflicted withthe underlying disorder. In certain embodiments, for prophylacticbenefit, the compositions are administered to a subject at risk ofdeveloping a particular disease, or to a subject reporting one or moreof the physiological symptoms of a disease, even though a diagnosis ofthis disease may not have been made.

A “therapeutic effect,” as that term is used herein, encompasses atherapeutic benefit and/or a prophylactic benefit as described above. Aprophylactic effect includes delaying or eliminating the appearance of adisease or condition, delaying or eliminating the onset of symptoms of adisease or condition, slowing, halting, or reversing the progression ofa disease or condition, or any combination thereof.

The term “co-administration,” “administered in combination with,” andtheir grammatical equivalents, as used herein, encompass administrationof two or more agents to an animal, including humans, so that bothagents and/or their metabolites are present in the subject at the sametime. Co-administration includes simultaneous administration in separatecompositions, administration at different times in separatecompositions, or administration in a composition in which both agentsare present.

“Pharmaceutically acceptable salt” includes both acid and base additionsalts.

“Pharmaceutically acceptable acid addition salt” refers to those saltswhich retain the biological effectiveness and properties of the freebases, which are not biologically or otherwise undesirable, and whichare formed with inorganic acids such as, but are not limited to,hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid,phosphoric acid and the like, and organic acids such as, but not limitedto, acetic acid, 2,2-dichloroacetic acid, adipic acid, alginic acid,ascorbic acid, aspartic acid, benzenesulfonic acid, benzoic acid,4-acetamidobenzoic acid, camphoric acid, camphor-10-sulfonic acid,capric acid, caproic acid, caprylic acid, carbonic acid, cinnamic acid,citric acid, cyclamic acid, dodecylsulfuric acid, ethane-1,2-disulfonicacid, ethanesulfonic acid, 2-hydroxyethanesulfonic acid, formic acid,fumaric acid, galactaric acid, gentisic acid, glucoheptonic acid,gluconic acid, glucuronic acid, glutamic acid, glutaric acid,2-oxo-glutaric acid, glycerophosphoric acid, glycolic acid, hippuricacid, isobutyric acid, lactic acid, lactobionic acid, lauric acid,maleic acid, malic acid, malonic acid, mandelic acid, methanesulfonicacid, mucic acid, naphthalene-1,5-disulfonic acid,naphthalene-2-sulfonic acid, 1-hydroxy-2-naphthoic acid, nicotinic acid,oleic acid, orotic acid, oxalic acid, palmitic acid, pamoic acid,propionic acid, pyroglutamic acid, pyruvic acid, salicylic acid,4-aminosalicylic acid, sebacic acid, stearic acid, succinic acid,tartaric acid, thiocyanic acid, p-toluenesulfonic acid, trifluoroaceticacid, undecylenic acid, and the like.

“Pharmaceutically acceptable base addition salt” refers to those saltswhich retain the biological effectiveness and properties of the freeacids, which are not biologically or otherwise undesirable. These saltsare prepared from addition of an inorganic base or an organic base tothe free acid. Salts derived from inorganic bases include, but are notlimited to, the sodium, potassium, lithium, ammonium, calcium,magnesium, iron, zinc, copper, manganese, aluminum salts and the like.Preferred inorganic salts are the ammonium, sodium, potassium, calcium,and magnesium salts. Salts derived from organic bases include, but arenot limited to, salts of primary, secondary, and tertiary amines,substituted amines including naturally occurring substituted amines,cyclic amines and basic ion exchange resins, such as ammonia,isopropylamine, trimethylamine, diethylamine, triethylamine,tripropylamine, diethanolamine, ethanolamine, deanol,2-dimethylaminoethanol, 2-diethylaminoethanol, dicyclohexylamine,lysine, arginine, histidine, caffeine, procaine, hydrabamine, choline,betaine, benethamine, benzathine, ethylenediamine, glucosamine,methylglucamine, theobromine, triethanolamine, tromethamine, purines,piperazine, piperidine, N-ethylpiperidine, polyamine resins and thelike. Particularly preferred organic bases are isopropylamine,diethylamine, ethanolamine, trimethylamine, dicyclohexylamine, cholineand caffeine.

The terms “antagonist” and “inhibitor” are used interchangeably, andthey refer to a compound having the ability to inhibit a biologicalfunction of a target protein, whether by inhibiting the activity orexpression of the protein, such as KRAS, HRAS or NRAS G12C. Accordingly,the terms “antagonist” and “inhibitors” are defined in the context ofthe biological role of the target protein. While preferred antagonistsherein specifically interact with (e.g. bind to) the target, compoundsthat inhibit a biological activity of the target protein by interactingwith other members of the signal transduction pathway of which thetarget protein is a member are also specifically included within thisdefinition. A preferred biological activity inhibited by an antagonistis associated with the development, growth, or spread of a tumor.

The term “agonist” as used herein refers to a compound having theability to initiate or enhance a biological function of a targetprotein, whether by inhibiting the activity or expression of the targetprotein. Accordingly, the term “agonist” is defined in the context ofthe biological role of the target polypeptide. While preferred agonistsherein specifically interact with (e.g. bind to) the target, compoundsthat initiate or enhance a biological activity of the target polypeptideby interacting with other members of the signal transduction pathway ofwhich the target polypeptide is a member are also specifically includedwithin this definition.

As used herein, “agent” or “biologically active agent” refers to abiological, pharmaceutical, or chemical compound or other moiety.Non-limiting examples include a simple or complex organic or inorganicmolecule, a peptide, a protein, an oligonucleotide, an antibody, anantibody derivative, antibody fragment, a vitamin derivative, acarbohydrate, a toxin, or a chemotherapeutic compound. Various compoundscan be synthesized, for example, small molecules and oligomers (e.g.,oligopeptides and oligonucleotides), and synthetic organic compoundsbased on various core structures. In addition, various natural sourcescan provide compounds for screening, such as plant or animal extracts,and the like.

“Signal transduction” is a process during which stimulatory orinhibitory signals are transmitted into and within a cell to elicit anintracellular response. A modulator of a signal transduction pathwayrefers to a compound which modulates the activity of one or morecellular proteins mapped to the same specific signal transductionpathway. A modulator may augment (agonist) or suppress (antagonist) theactivity of a signaling molecule.

An “anti-cancer agent”, “anti-tumor agent” or “chemotherapeutic agent”refers to any agent useful in the treatment of a neoplastic condition.One class of anti-cancer agents comprises chemotherapeutic agents.“Chemotherapy” means the administration of one or more chemotherapeuticdrugs and/or other agents to a cancer patient by various methods,including intravenous, oral, intramuscular, intraperitoneal,intravesical, subcutaneous, transdermal, buccal, or inhalation or in theform of a suppository.

The term “cell proliferation” refers to a phenomenon by which the cellnumber has changed as a result of division. This term also encompassescell growth by which the cell morphology has changed (e.g., increased insize) consistent with a proliferative signal.

The term “selective inhibition” or “selectively inhibit” refers to abiologically active agent refers to the agent's ability topreferentially reduce the target signaling activity as compared tooff-target signaling activity, via direct or indirect interaction withthe target.

“Subject” refers to an animal, such as a mammal, for example a human.The methods described herein can be useful in both human therapeuticsand veterinary applications. In some embodiments, the subject is amammal, and in some embodiments, the subject is human.

“Mammal” includes humans and both domestic animals such as laboratoryanimals and household pets (e.g., cats, dogs, swine, cattle, sheep,goats, horses, rabbits), and non-domestic animals such as wildlife andthe like.

“Radiation therapy” means exposing a subject, using routine methods andcompositions known to the practitioner, to radiation emitters such asalpha-particle emitting radionuclides (e.g., actinium and thoriumradionuclides), low linear energy transfer (LET) radiation emitters(i.e. beta emitters), conversion electron emitters (e.g. strontium-89and samarium-153-EDTMP, or high-energy radiation, including withoutlimitation x-rays, gamma rays, and neutrons.

An “anti-cancer agent”, “anti-tumor agent” or “chemotherapeutic agent”refers to any agent useful in the treatment of a neoplastic condition.One class of anti-cancer agents comprises chemotherapeutic agents.“Chemotherapy” means the administration of one or more chemotherapeuticdrugs and/or other agents to a cancer patient by various methods,including intravenous, oral, intramuscular, intraperitoneal,intravesical, subcutaneous, transdermal, buccal, or inhalation or in theform of a suppository.

“Prodrug” is meant to indicate a compound that may be converted underphysiological conditions or by solvolysis to a biologically activecompound described herein (e.g., compound of structure (I)). Thus, theterm “prodrug” refers to a precursor of a biologically active compoundthat is pharmaceutically acceptable. In some aspects, a prodrug isinactive when administered to a subject, but is converted in vivo to anactive compound, for example, by hydrolysis. The prodrug compound oftenoffers advantages of solubility, tissue compatibility or delayed releasein a mammalian organism (see, e.g., Bundgard, H., Design of Prodrugs(1985), pp. 7-9, 21-24 (Elsevier, Amsterdam). A discussion of prodrugsis provided in Higuchi, T., et al., “Pro-drugs as Novel DeliverySystems,” A.C.S. Symposium Series, Vol. 14, and in BioreversibleCarriers in Drug Design, ed. Edward B. Roche, American PharmaceuticalAssociation and Pergamon Press, 1987, both of which are incorporated infull by reference herein. The term “prodrug” is also meant to includeany covalently bonded carriers, which release the active compound invivo when such prodrug is administered to a mammalian subject. Prodrugsof an active compound, as described herein, are typically prepared bymodifying functional groups present in the active compound in such a waythat the modifications are cleaved, either in routine manipulation or invivo, to the parent active compound. Prodrugs include compounds whereina hydroxy, amino or mercapto group is bonded to any group that, when theprodrug of the active compound is administered to a mammalian subject,cleaves to form a free hydroxy, free amino or free mercapto group,respectively. Examples of prodrugs include, but are not limited to,acetate, formate and benzoate derivatives of a hydroxy functional group,or acetamide, formamide and benzamide derivatives of an amine functionalgroup in the active compound and the like.

In some embodiments, prodrugs include compounds of structure (I) havinga phosphate, phosphoalkoxy, ester or boronic ester substituent. Withoutbeing bound by theory, it is believed that such substituents areconverted to a hydroxyl group under physiological conditions.Accordingly, embodiments include any of the compounds disclosed herein,wherein a hydroxyl group has been replaced with a phosphate,phosphoalkoxy, ester or boronic ester group, for example a phosphate orphosphoalkoxy group. For example, in some embodiments a hydroxyl groupon the R¹ moiety is replaced with a phosphate, phosphoalkoxy, ester orboronic ester group, for example a phosphate or alkoxy phosphate group.Exemplary prodrugs of certain embodiments thus include compounds havingone of the following R¹ moieties:

wherein each R′ is independently H or an optional substituent, and n is1, 2, 3 or 4.

The term “in vivo” refers to an event that takes place in a subject'sbody.

Embodiments of the invention disclosed herein are also meant toencompass all pharmaceutically acceptable compounds of structure (I)being isotopically-labelled by having one or more atoms replaced by anatom having a different atomic mass or mass number. Examples of isotopesthat can be incorporated into the disclosed compounds include isotopesof hydrogen, carbon, nitrogen, oxygen, phosphorous, fluorine, chlorine,and iodine, such as ²H, ³H, ¹¹C, ¹³C, ¹⁴C, ¹³N, ¹⁵N, ¹⁵O, ¹⁷O, ¹⁸O, ³¹P,³²P, ³⁵S, ¹⁸F, ³⁶Cl, ¹²³I, and ¹²⁵I, respectively. These radiolabeledcompounds could be useful to help determine or measure the effectivenessof the compounds, by characterizing, for example, the site or mode ofaction, or binding affinity to pharmacologically important site ofaction. Certain isotopically-labeled compounds of structure (I), forexample, those incorporating a radioactive isotope, are useful in drugand/or substrate tissue distribution studies. The radioactive isotopestritium, i.e. ³H, and carbon-14, i.e. ¹⁴C, are particularly useful forthis purpose in view of their ease of incorporation and ready means ofdetection.

Substitution with heavier isotopes such as deuterium, i.e. ²H, mayafford certain therapeutic advantages resulting from greater metabolicstability, for example, increased in vivo half-life or reduced dosagerequirements, and hence are preferred in some circumstances.

Substitution with positron emitting isotopes, such as ¹¹C, ¹⁸F, ¹⁵O and¹³N, can be useful in Positron Emission Topography (PET) studies forexamining substrate receptor occupancy. Isotopically-labeled compoundsof structure (I) can generally be prepared by conventional techniquesknown to those skilled in the art or by processes analogous to thosedescribed in the Preparations and Examples as set out below using anappropriate isotopically-labeled reagent in place of the non-labeledreagent previously employed.

Certain embodiments are also meant to encompass the in vivo metabolicproducts of the disclosed compounds. Such products may result from, forexample, the oxidation, reduction, hydrolysis, amidation,esterification, and the like of the administered compound, primarily dueto enzymatic processes. Accordingly, embodiments include compoundsproduced by a process comprising administering a compound of thisinvention to a mammal for a period of time sufficient to yield ametabolic product thereof. Such products are typically identified byadministering a radiolabeled compound of the invention in a detectabledose to an animal, such as rat, mouse, guinea pig, monkey, or to human,allowing sufficient time for metabolism to occur, and isolating itsconversion products from the urine, blood or other biological samples.

“Stable compound” and “stable structure” are meant to indicate acompound that is sufficiently robust to survive isolation to a usefuldegree of purity from a reaction mixture, and formulation into anefficacious therapeutic agent.

Often crystallizations produce a solvate of the compound of theinvention. As used herein, the term “solvate” refers to an aggregatethat comprises one or more molecules of a compound of the invention withone or more molecules of solvent. In some embodiments, the solvent iswater, in which case the solvate is a hydrate. Alternatively, in otherembodiments, the solvent is an organic solvent. Thus, the compounds ofthe present invention may exist as a hydrate, including a monohydrate,dihydrate, hemihydrate, sesquihydrate, trihydrate, tetrahydrate and thelike, as well as the corresponding solvated forms. In some aspects, thecompound of the invention is a true solvate, while in other cases, thecompound of the invention merely retains adventitious water or is amixture of water plus some adventitious solvent.

“Optional” or “optionally” means that the subsequently described eventof circumstances may or may not occur, and that the description includesinstances where said event or circumstance occurs and instances in whichit does not. For example, “optionally substituted aryl” means that thearyl radical may or may not be substituted and that the descriptionincludes both substituted aryl radicals and aryl radicals having nosubstitution.

A “pharmaceutical composition” refers to a formulation of a compound ofthe invention and a medium generally accepted in the art for thedelivery of the biologically active compound to mammals, e.g., humans.Such a medium includes all pharmaceutically acceptable carriers,diluents or excipients therefor.

“Pharmaceutically acceptable carrier, diluent or excipient” includeswithout limitation any adjuvant, carrier, excipient, glidant, sweeteningagent, diluent, preservative, dye/colorant, flavor enhancer, surfactant,wetting agent, dispersing agent, suspending agent, stabilizer, isotonicagent, solvent, or emulsifier which has been approved by the UnitedStates Food and Drug Administration as being acceptable for use inhumans or domestic animals.

The compounds of the invention (i.e., compounds of structure (I)), ortheir pharmaceutically acceptable salts may contain one or moreasymmetric centers and may thus give rise to enantiomers, diastereomers,and other stereoisomeric forms that are defined, in terms of absolutestereochemistry, as (R)- or (S)- or, as (D)- or (L)- for amino acids.Embodiments thus include all such possible isomers, as well as theirracemic and optically pure forms. Optically active (+) and (−), (R)- and(S)-, or (D)- and (L)-isomers may be prepared using chiral synthons orchiral reagents, or resolved using conventional techniques, for example,chromatography and fractional crystallization. Conventional techniquesfor the preparation/isolation of individual enantiomers include chiralsynthesis from a suitable optically pure precursor or resolution of theracemate (or the racemate of a salt or derivative) using, for example,chiral high pressure liquid chromatography (HPLC). When the compoundsdescribed herein contain olefinic double bonds or other centres ofgeometric asymmetry, and unless specified otherwise, it is intended thatthe compounds include both E and Z geometric isomers. Likewise, alltautomeric forms are also intended to be included.

Embodiments of the present invention include all manner of rotamers andconformationally restricted states of a compound of the invention.Atropisomers, which are stereoisomers arising because of hinderedrotation about a single bond, where energy differences due to stericstrain or other contributors create a barrier to rotation that is highenough to allow for isolation of individual conformers, are alsoincluded. As an example, certain compounds of the invention may exist asmixtures of atropisomers or purified or enriched for the presence of oneatropisomer. Atropisomers of the present invention are isolated and/orpurified using techniques known in the art, such chiral chromatography.Non-limiting examples of compounds which exist as atropisomers includethe following compound:

A “stereoisomer” refers to a compound made up of the same atoms bondedby the same bonds but having different three-dimensional structures,which are not interchangeable. The present invention contemplatesvarious stereoisomers and mixtures thereof and includes “enantiomers”,which refers to two stereoisomers whose molecules are nonsuperimposablemirror images of one another.

A “tautomer” refers to a proton shift from one atom of a molecule toanother atom of the same molecule. Embodiments thus include tautomers ofthe disclosed compounds.

The chemical naming protocol and structure diagrams used herein are amodified form of the I.U.P.A.C. nomenclature system, using the ACD/NameVersion 9.07 software program and/or ChemDraw Ultra Version 11.0.1software naming program (CambridgeSoft). For complex chemical namesemployed herein, a substituent group is typically named before the groupto which it attaches. For example, cyclopropylethyl comprises an ethylbackbone with a cyclopropyl substituent. Except as described below, allbonds are identified in the chemical structure diagrams herein, exceptfor all bonds on some carbon atoms, which are assumed to be bonded tosufficient hydrogen atoms to complete the valency.

Compounds

In an aspect, the invention provides compounds which are capable ofselectively binding to and/or modulating a G12C mutant KRAS, HRAS orNRAS protein. The compounds may modulate the G12C mutant KRAS, HRAS orNRAS protein by reaction with an amino acid. While not wishing to bebound by theory, the present applicants believe that, in someembodiments, the compounds of the invention selectively react with theG12C mutant KRAS, HRAS or NRAS proteins by forming a covalent bond withthe cysteine at the 12 position of a G12C mutant KRAS, HRAS or NRASprotein. By binding to the Cystine 12, the compounds of the inventionmay lock the switch II of the G12C mutant KRAS, HRAS or NRAS into aninactive stage. This inactive stage may be distinct from those observedfor GTP and GDP bound KRAS, HRAS or NRAS. Some compounds of theinvention may also be able to perturb the switch I conformation. Somecompounds of the invention may favor the binding of the bound KRAS, HRASor NRAS to GDP rather than GTP and therefore sequester the KRAS, HRAS orNRAS into an inactive KRAS, HRAS or NRAS GDP state. Because effectorbinding to KRAS, HRAS or NRAS is highly sensitive to the conformation ofswitch I and II, the irreversible binding of these compounds may disruptKRAS, HRAS or NRAS downstream signaling.

As noted above, in one embodiment of the present invention, compoundshaving activity as modulators of a G12C mutant KRAS, HRAS or NRASprotein are provided, the compounds have the following structure (I):

or a pharmaceutically acceptable salt, stereoisomer or prodrug thereof,wherein:

A is a carbocyclic, heterocyclic or heteroaryl ring;

G¹ and G² are each independently N or CH;

L¹ is a bond or NR⁵;

L² is a bond or alkylene;

R¹ is aryl or heteroaryl;

R^(2a), R^(2b) and R^(2c) are each independently H, amino, halo,hydroxyl, cyano, C₁-C₆ alkyl, C₁-C₆ alkyl amino, C₁-C₆ haloalkyl, C₁-C₆alkoxy, C₁-C₆ haloalkoxy; C₃-C₈ cycloalkyl, heterocyclylalkyl, C₁-C₆alkynyl, C₁-C₆ alkenyl, aminylalkyl, alkylaminylalkyl, cyanoalkyl,carboxyalkyl, aminylcarbonylalkyl, aminylcarbonyl, heteroaryl or aryl;

R^(3a) and R^(3b) are, at each occurrence, independently H, —OH, —NH₂,—CO₂H, halo, cyano, C₁-C₆ alkyl, C₁-C₆ haloalkyl, C₁-C₆ haloalkoxy,C₃-C₈ cycloalkyl, heterocyclylalkyl, C₁-C₆ alkynyl, hydroxylalkly,alkoxyalkyl, aminylalkyl, alkylaminylalkyl, cyanoalkyl, carboxyalkyl,aminylcarbonylalkyl or aminylcarbonyl; or R^(3a) and R^(3b) join to forma carbocyclic or heterocyclic ring; or R^(3a) is H, —OH, —NH₂, —CO₂H,halo, cyano, C₁-C₆ alkyl, C₁-C₆ haloalkyl, C₁-C₆ haloalkoxy, C₃-C₈cycloalkyl, heterocyclylalkyl, C₁-C₆ alkynyl, hydroxylalkly,alkoxyalkyl, aminylalkyl, alkylaminylalkyl, cyanoalkyl, carboxyalkyl,aminylcarbonylalkyl or aminylcarbonyl, and R^(3b) joins with R^(4b) toform a carbocyclic or heterocyclic ring;

R^(4a) and R^(4b) are, at each occurrence, independently H, —OH, —NH₂,—CO₂H, halo, cyano, C₁-C₆ alkyl, C₁-C₆ haloalkyl, C₁-C₆ haloalkoxy,C₃-C₈ cycloalkyl, heterocyclylalkyl, C₁-C₆ alkynyl, hydroxylalkly,alkoxyalkyl, aminylalkyl, alkylaminylalkyl, cyanoalkyl, carboxyalkyl,aminylcarbonylalkyl or aminylcarbonyl; or R^(4a) and R^(4b) join to forma carbocyclic or heterocyclic ring; or R^(4a) is H, —OH, —NH₂, —CO₂H,halo, cyano, C₁-C₆ alkyl, C₁-C₆ haloalkyl, C₁-C₆ haloalkoxy, C₃-C₈cycloalkyl, heterocyclylalkyl, C₁-C₆ alkynyl, hydroxylalkly,alkoxyalkyl, aminylalkyl, alkylaminylalkyl, cyanoalkyl, carboxyalkyl,aminylcarbonylalkyl or aminylcarbonyl, and R^(4b) joins with R^(3b) toform a carbocyclic or heterocyclic ring;

R⁵ is, at each occurrence, independently H, C₁-C₆ alkyl, C₃-C₈cycloalkyl or heterocycloalkyl;

m¹ and m² are each independently 1, 2 or 3; and

E is an electrophilic moiety capable of forming a covalent bond with thecysteine residue at position 12 of a KRAS, HRAS or NRAS G12C mutantprotein.

In other embodiments of compounds of structure (I):

A is a carbocyclic, heterocyclic or heteroaryl ring;

G¹ and G² are each independently N or CH;

L¹ is a bond or NR⁵;

L² is a bond or alkylene;

R¹ is aryl or heteroaryl;

R^(2a), R^(2b) and R^(2c) are each independently H, amino, halo,hydroxyl, cyano, C₁-C₆ alkyl, C₁-C₆ alkyl amino, C₁-C₆ haloalkyl, C₁-C₆alkoxy, C₃-C₈ cycloalkyl, heteroaryl or aryl;

R^(3a) and R^(3b) are, at each occurrence, independently H, —OH, —NH₂,—CO₂H, halo, cyano, C₁-C₆ alkyl, C₁-C₆ alkynyl, hydroxylalkly,alkoxyalkyl, aminylalkyl, alkylaminylalkyl, cyanoalkyl, carboxyalkyl,aminylcarbonylalkyl or aminylcarbonyl; or R^(3a) and R^(3b) join to forma carbocyclic or heterocyclic ring; or R^(3a) is H, —OH, —NH₂, —CO₂H,halo, cyano, C₁-C₆ alkyl, C₁-C₆ alkynyl, hydroxylalkly, alkoxyalkyl,aminylalkyl, alkylaminylalkyl, cyanoalkyl, carboxyalkyl,aminylcarbonylalkyl or aminylcarbonyl, and R^(3b) joins with R^(4b) toform a carbocyclic or heterocyclic ring;

R^(4a) and R^(4b) are, at each occurrence, independently H, —OH, —NH₂,—CO₂H, halo, cyano, C₁-C₆ alkyl, C₁-C₆ alkynyl, hydroxylalkly,alkoxyalkyl, aminylalkyl, alkylaminylalkyl, cyanoalkyl, carboxyalkyl,aminylcarbonylalkyl or aminylcarbonyl; or R^(4a) and R^(4b) join to forma carbocyclic or heterocyclic ring; or R^(4a) is H, —OH, —NH₂, —CO₂H,halo, cyano, C₁-C₆ alkyl, C₁-C₆ alkynyl, hydroxylalkly, alkoxyalkyl,aminylalkyl, alkylaminylalkyl, cyanoalkyl, carboxyalkyl,aminylcarbonylalkyl or aminylcarbonyl, and R^(4b) joins with R^(3b) toform a carbocyclic or heterocyclic ring;

R⁵ is, at each occurrence, independently H, C₁-C₆ alkyl, C₃-C₈cycloalkyl or heterocycloalkyl;

m¹ and m² are each independently 1, 2 or 3; and

E is an electrophilic moiety capable of forming a covalent bond with thecysteine residue at position 12 of a KRAS, HRAS or NRAS G12C mutantprotein.

The structure of E is not particularly limited provided it is capable offorming a covalent bond with a nucleophile, such as the cysteine residueat position 12 of a KRAS, HRAS or NRAS G12C mutant protein. Accordingly,moieties which are capable of reaction with (e.g., by covalent bondformation) a nucleophile are preferred. In certain embodiments, E iscapable of reacting in a conjugate addition manner (e.g., 1,4-conjugateaddition) with an appropriately reactive nucleophile. In someembodiments, E comprises conjugated pi bonds such that delocalization ofelectrons results in at least one atom (e.g., a carbon atom) having apositive charge, partial positive charge or a polarized bond. In otherembodiments, E comprises one or more bonds wherein the electronegativityof the two atoms forming the bonds is sufficiently different such that apartial positive charge (e.g., by polarization of the bond) resides onone of the atoms, for example on a carbon atom. E moieties comprisingcarbon-halogen bonds, carbon-oxygen bonds or carbon bonds to variousleaving groups known in the art are examples of such E moieties.

In certain embodiments of the foregoing, E has the following structure:

wherein:

represents a double or triple bond;

Q is —C(═O)—, —C(═NR^(8′))—, —NR⁸C(═O)—, —S(═O)₂— or —NR⁸S(═O)₂—;

R⁸ is H, C₁-C₆ alkyl, hydroxylalkyl, aminoalkyl, alkoxyalkyl,aminylalkyl, alkylaminylalkyl, cyanoalkyl, carboxyalkyl,aminylcarbonylalkyl, C₃-C₈ cycloalkyl or heterocycloalkyl;

R^(8′) is H, —OH, —CN or C₁-C₆ alkyl; and

when

is a double bond then R⁹ and R¹⁰ are each independently H, halo, cyano,carboxyl, C₁-C₆ alkyl, alkoxycarbonyl, aminylalkyl, alkylaminylalkyl,aryl, heterocyclyl, heterocyclylalkyl, heteroaryl or hydroxylalkyl, orR⁹ and R¹⁰ join to form a carbocyclic, heterocyclic or heteroaryl ring;

when

is a triple bond; then R⁹ is absent and R¹⁰ is H, C₁-C₆alkyl,aminylalkyl, alkylaminylalkyl or hydroxylalkyl.

In certain embodiments when

is a double bond then R⁹ and R¹⁰ are each independently H, cyano,C₁-C₆alkyl, aminylalkyl, alkylaminylalkyl, or hydroxylalkyl or R⁹ andR¹⁰ join to form a carbocyclic or heterocyclic ring.

In some of the foregoing embodiments, Q is —C(═O)—, —NR⁸C(═O)—, —S(═O)₂—or —NR⁸S(═O)₂—.

In some other of the foregoing embodiments, Q is —C(═NR^(8′))—, whereinR^(8′) is H, —OH, —CN or C₁-C₆alkyl. For example, in some embodimentsR^(8′) is H. In other embodiments, R^(8′) is —CN. In other embodiments,R^(8′) is —OH.

Accordingly, in some embodiments, the compound has the followingstructure (I′a):

wherein:

represents a double or triple bond;

Q is —C(═O)—, —C(═NR^(8′))—, —NR⁸C(═O)—, —S(═O)₂— or —NR⁸S(═O)₂—;

R⁸ is H, C₁-C₆ alkyl, hydroxylalkyl, aminoalkyl, alkoxyalkyl,aminylalkyl, alkylaminylalkyl, cyanoalkyl, carboxyalkyl,aminylcarbonylalkyl, C₃-C₈ cycloalkyl or heterocycloalkyl;

R^(8′) is H, —OH, —CN or C₁-C₆ alkyl;

when

is a double on then R⁹ and R¹⁰ are each independently H, halo, cyano,carboxyl, C₁-C₆ alkyl, alkoxycarbonyl, aminylalkyl, alkylaminylalkyl,aryl, heterocyclyl, heterocyclylalkyl, heteroaryl or hydroxylalkyl, orR⁹ and R¹⁰ join to form a carbocyclic, heterocyclic or heteroaryl ring;and

when

is a triple bond then R⁹ is absent and R¹⁰ is H, C₁-C₆ alkyl,aminylalkyl, alkylaminylalkyl or hydroxylalkyl.

In other embodiments, the compound has one of the following structures(I′b), (I′c), (I′d) or (I′e):

In some of the foregoing embodiments, A is a 5, 6 or 7-memberedheterocyclic or heteroaryl ring. In some embodiments, A is a 5-memberedheterocyclic ring. In other embodiments, A is a 5-membered heteroarylring. In some embodiments, A is a 6-membered heterocyclic ring. In otherembodiments, A is a 6-membered heteroaryl ring. In some embodiments, Ais a 7-membered heterocyclic ring. In other embodiments, A is a7-membered heteroaryl ring. In some embodiments, A has one of thefollowing structures:

wherein:

R⁶ is, at each occurrence, independently H, halo, amino, cyano, arayl,heteroaryl, C₁-C₆ alkyl, C₃-C₈ cycloalkyl, heterocycloalkyl,hydroxylalkly, alkoxyalkyl, aminylalkyl, alkylaminylalkyl, cyanoalkyl,carboxyalkyl, aminylcarbonylalkyl, arylalkyl or heteroarylalkyl;

X is O or CH₂; and

n is 0, 1 or 2.

For example, in some different embodiments, A has one of the followingstructures:

wherein:

R⁶ is, at each occurrence, independently H, C₁-C₆ alkyl, C₃-C₈cycloalkyl, heterocycloalkyl, hydroxylalkly, alkoxyalkyl, aminylalkyl,alkylaminylalkyl, cyanoalkyl, carboxyalkyl, aminylcarbonylalkyl,arylalkyl or heteroarylalkyl;

X is O or CH₂; and

n is 0, 1 or 2.

Without wishing to be bound by theory, Applicants believe correctselection of the R¹ substituent may play a part in the compounds'inhibitory activity (e.g., against KRAS, HRAS or NRAS G12C). In someembodiments, R¹ is aryl or heterocyclyl (e.g., heteroaryl or aliphaticheterocyclyl), each of which is optionally substituted with one or moresubstituents. In some embodiments, R¹ is capable of reversibleinteraction with KRAS, HRAS or NRAS G12C mutant protein. In someembodiments R¹ has high affinity towards KRAS, HRAS or NRAS and ishighly specific towards G12C KRAS, HRAS or NRAS. In some embodiments R¹is capable of hydrophobic interaction with KRAS, HRAS or NRAS G12C. Insome embodiments R¹ is able to form hydrogen bonds with various residuesof G12C KRAS, HRAS or NRAS protein.

In any of the foregoing embodiments, R¹ is aryl. For example in someembodiments R¹ is phenyl, and in other embodiments R¹ is naphthyl. R¹ issubstituted or unsubstituted. In some specific embodiments, R¹ issubstituted with one or more substituents. In various embodiments, R¹ issubstituted with halo, amino hydroxyl, C₁-C₆ alkyl, cyano, C₁-C₆haloalkyl, C₁-C₆ alkoxy, alkylaminyl, cycloalkyl, heterocyclylalkyl,aryl, heteroaryl, boronic acid, —OC(═O)R, phosphate, phosphoalkoxy orC₁-C₆ alkylcarbonyloxy, or combinations thereof, wherein R is C₁-C₆alkyl. For example, in some embodiments R¹ is substituted with halo,hydroxyl, C₁-C₆ alkyl, C₁-C₆ haloalkyl, C₁-C₆ alkoxy or C₁-C₆alkylcarbonyloxy, or combinations thereof. In still other embodiments,R¹ is substituted with fluoro, chloro, hydroxyl, methyl, isopropyl,cyclopropyl, trifluoromethyl or methoxy, or combinations thereof. Insome even more embodiments, R¹ is substituted with fluoro or hydroxyl,or combinations thereof.

In some different embodiments of the foregoing compounds, R¹ isheteroaryl, for example a heteroaryl comprising nitrogen. In otherembodiments, R¹ is indazolyl, indolyl, benzoimidazole, benzotriazole orquinolinyl, for example indazolyl or quinolinyl. In more embodiments, R¹is heteroaryl which is substituted with one or more substituents. Forexample, in certain embodiments, R¹ is substituted with hydroxyl, haloor C₁-C₆ alkyl, or combinations thereof, for example hydroxyl or C₁-C₆alkyl, or both.

In certain embodiments, R¹ has one of the following structures:

In some more specific embodiments, R¹ has one of the followingstructures:

In some embodiments, R^(2a), R^(2b) and/or R^(2c) are CF₃. In some ofthe foregoing embodiments R^(2c) is H. In some embodiments, R^(2a) andR^(2b) are each independently halo, haloalkyl, alkyl or alkoxy. In otherof any of the foregoing embodiments, R^(2a) and R^(2b) are each halo.For example, in some embodiments R^(2a) is fluoro, and in otherembodiments, R^(2b) is chloro. In other embodiments, R^(2a) is fluoro,chloro or methoxy. In different embodiments, R^(2b) is chloro, fluoro orCF₃.

In some more specific embodiments, the compounds have the followingstructure (I′f):

For example, in even further different embodiments, the compounds havethe following structure (I′g):

In yet more of any of the foregoing embodiments, E has the followingstructure:

wherein:

Q is —C(═O)—, —C(═NR^(8′))—, —NR⁸C(═O)—, —S(═O)₂— or —NR⁸S(═O)₂—;

R⁸ is H, C₁-C₆alkyl or hydroxylalkyl;

R^(8′) is H, —OH, —CN or C₁-C₆alkyl; and

R⁹ and R¹⁰ are each independently H, halo, cyano, carboxyl, C₁-C₆ alkyl,alkoxycarbonyl, aminylalkyl, alkylaminylalkyl, aryl, heterocyclyl,heterocyclylalkyl, heteroaryl or hydroxylalkyl, or R⁹ and R¹⁰ join toform a carbocyclic, heterocyclic or heteroaryl ring.

In still other of any of the foregoing embodiments, E has the followingstructure:

wherein:

Q is —C(═O)—, —NR⁸C(═O)—, —S(═O)₂— or —NR⁸S(═O)₂—;

R⁸ is H, C₁-C₆alkyl or hydroxylalkyl; and

R¹⁰ is H, C₁-C₆alkyl, aminylalkyl, alkylaminylalkyl or hydroxylalkyl.

The Q moiety is typically selected to optimize the reactivity (i.e.,electrophilicity) of E. In some of the foregoing embodiments Q is—C(═O)—, —NR⁸C(═O)—, —S(═O)₂— or —NR⁸S(═O)₂—. In certain of theforegoing embodiments, Q is —C(═O)—. In other embodiments, Q is—S(═O)₂—. In still more embodiments, Q is —NR⁸C(═O)—. In still moredifferent embodiments, Q is —NR⁸S(═O)₂—.

In some other of the foregoing embodiments, Q is —C(═NR^(8′))—, whereinR^(8′) is H, —OH, —CN or C₁-C₆alkyl. For example, in some embodimentsR^(8′) is H. In other embodiments, R^(8′) is —CN. In other embodiments,R^(8′) is —OH.

In some of the foregoing embodiments, R⁸ is H. In other of theseembodiments, R⁸ is hydroxylalkyl, for example in some embodiments thehydroxylalkyl is 2-hydroxylalkyl.

In some of any one of the foregoing embodiments, at least one of R⁹ orR¹⁰ is H. For example, in some embodiments each of R⁹ and R¹⁰ are H.

In other of the foregoing embodiments, R¹⁰ is alkylaminylalkyl. In someof these embodiments, R¹⁰ has the following structure:

In other embodiments, R¹⁰ is hydroxylalkyl, such as 2-hydroxylalkyl.

In some other different embodiments of the foregoing embodiments, R⁹ andR¹⁰ join to form a carbocyclic ring. For example, in some of theseembodiments the carbocyclic ring is a cyclopentene, cyclohexene orphenyl ring. In other embodiments, the carbocyclic ring is acyclopentene or cyclohexene ring. In other embodiments, the carbocyclicring is a phenyl ring, for example a phenyl ring having the followingstructure:

In some of any of the foregoing embodiments E is an electrophile capableof bonding with a KRAS, HRAS or NRAS protein comprising G12C mutation.In some embodiments, the electrophile E is capable of forming anirreversible covalent bond with a G12C mutant KRAS, HRAS or NRASprotein. In some cases, the electrophile E may bind with the cysteineresidue at the position 12 of a G12C mutant KRAS, HRAS or NRAS protein.In various embodiments of any of the foregoing, E has one of thefollowing structures:

In some embodiments, E has one of the following structures:

In other embodiments of any of the foregoing, E has one of the followingstructures:

In different embodiments, E has one of the following structures:

In some cases E has one of the following structures:

wherein:

R⁸ is H or C₁-C₆alkyl;

R⁹ is H, cyano or C₁-C₆alkyl, or R⁹ joins with R¹⁰ to form a carbocycle;

R¹⁰ is H or C₁-C₆alkyl or R¹⁰ joins with R⁹ to form a carbocycle and

R^(10a) is H or C₁-C₆alkyl.

In some embodiments E is

In some embodiments E is

In some embodiments E is

In some of any of the foregoing embodiments, L¹ is a bond. In otherembodiments, L¹ is NR⁵. For example, in some of these embodiments, R⁵ isC₁-C₆alkyl. In other embodiments, L¹ is NH.

L² can be selected to provide proper spacing and/or orientation for theE group to form a bond with the KRAS, HRAS or NRAS protein. In some ofthe foregoing embodiments, L² is a bond. In other of the foregoingembodiments, L² is alkylene. In some embodiments, the alkylene issubstituted. In other embodiments the alkylene is unsubstituted. Forexample, in some embodiments L² is CH₂ or CH₂CH₂.

In certain embodiments, R^(3a) and R^(3b) are, at each occurrence,independently H, —OH, —NH₂, —CO₂H, halo, cyano, hydroxylalkly,aminylalkyl, cyanoalkyl, carboxyalkyl or aminylcarbonyl, and R^(4a) andR^(4b) are, at each occurrence, independently H, —OH, —NH₂, —CO₂H, halo,cyano, hydroxylalkly, aminylalkyl, cyanoalkyl, carboxyalkyl oraminylcarbonyl.

In other of the foregoing embodiments, R^(3a) and R^(4a) are, at eachoccurrence, independently H, —OH, hydroxylalkly, cyano, oraminylcarbonyl and R^(3b) and R^(4b) are H.

In certain other embodiments, R^(3a) and R^(4a) are H and R^(3b) andR^(4b) are, at each occurrence, independently H, —OH, —NH₂, —CO₂H, halo,cyano, hydroxylalkly, aminylalkyl, cyanoalkyl, carboxyalkyl oraminylcarbonyl.

In any of the foregoing embodiments, at least one of R^(3a), R^(3b),R^(4a) or R^(4b) is H. In some embodiments, each of R^(3a), R^(3b),R^(4a) and R^(4b) are H.

In other of the foregoing embodiments, R^(3a) and R^(4a) are, at eachoccurrence, independently H or C₁-C₆ alkyl. In some embodiments, atleast one of R^(3a), R^(4a), R^(3b) and R^(4b) is independently C₁-C₆alkyl, such as methyl. In some embodiments, one occurrence of R^(3a) isC₁-C₆ alkyl, such as methyl, and each remaining R^(3a) and each R^(4a)is H. In some other embodiments, two occurrences of R^(3a) are C₁-C₆alkyl, such as methyl, and each remaining R^(3a) and each R^(4a) is H.In some other embodiments, one occurrence of R^(3a) and one occurrenceof R^(4a) is independently C₁-C₆ alkyl, such as methyl, and eachremaining R^(3a) and R^(4a) are each H.

In some embodiments, R^(3a) is —OH, —NH₂, —CO₂H, halo, cyano,hydroxylalkly, aminylalkyl, cyanoalkyl, carboxyalkyl or aminylcarbonyl,and R^(3b), R^(4a) and R^(4b) are H.

In other embodiments, R^(4a) is —OH, —NH₂, —CO₂H, halo, cyano,hydroxylalkly, aminylalkyl, cyanoalkyl, carboxyalkyl or aminylcarbonyl,and R^(3a), R^(3b) and R^(4b) are H.

In other embodiments, R^(3a) is H, —OH, —NH₂, —CO₂H, halo, cyano,hydroxylalkly, aminylalkyl, cyanoalkyl, carboxyalkyl or aminylcarbonyl,and R^(3b) joins with R^(4b) to form a carbocyclic or heterocyclic ring;

In still more embodiments, R^(4a) is H, —OH, —NH₂, —CO₂H, halo, cyano,hydroxylalkly, aminylalkyl, cyanoalkyl, carboxyalkyl or aminylcarbonyl,and R^(4b) joins with R^(3b) to form a carbocyclic or heterocyclic ring.

In other embodiments, R^(3a) and R^(3b) join to form a carbocyclic orheterocyclic ring. In other embodiments, R^(4a) and R^(4b) join to forma carbocyclic or heterocyclic ring.

In still other embodiments, R^(3a) or R^(4a) is aminylcarbonyl. Forexample, in certain embodiments, the aminylcarbonyl is

In other embodiments, R^(3a) or R^(4a) is cyano. In other embodiments,R^(3a) or R^(4a) is —OH. In other embodiments, R^(3a) or R^(4a) ishydroxylalkyl, for example hydroxylmethyl.

In some embodiments of any of the foregoing compounds, R¹ is aryl orheteroaryl and R^(2a), R^(2b) and R^(2c) are independently selected fromH and halo, for example in some further embodiments R¹ is aryl orheteroaryl and R^(2a) and R^(2b) are independently selected from halo,such as chloro and fluoro, and R^(2c) is H. In some embodiments, R¹ isaryl or heteroaryl, R^(2a) is chloro, R^(2b) is fluoro and R^(2c) is H.In other embodiments R¹ is aryl or heteroaryl, one of R^(2a) or R^(2b)is halo, such as chloro or fluoro, and the other one of R^(2a) or R^(2b)is H.

In some embodiments of any of the compounds described herein, C₁-C₆haloalkyl is CF₃ (e.g., when one or more of R^(2a), R^(2b) or R^(2c) isC₁-C₆ haloalkyl).

In some embodiments m¹ is 1. In other embodiments m¹ is 2. In still moreembodiments, m¹ is 3. In different embodiments, m² is 1. In some otherembodiments, m² is 2. In yet still more embodiments, m² is 3.

In some other particular embodiments of any of the foregoing compounds,m¹ is 1, and m² is 1. In other embodiments, m¹ is 1 and, m² is 2. Instill other embodiments m¹ is 2, and m² is 2. In more embodiments, m¹ is1, and m² is 3.

In any of the foregoing embodiments, G¹ and G² are each independentlyselected from N and CH. In some embodiments, at least one of G¹ or G² isN. In some embodiments, each of G¹ and G² are N. In some embodiments,each of G¹ and G² are N and m¹ and m² are each 2. In some otherembodiments, at least one of G¹ or G² is CH. In other embodiments, eachof G¹ and G² are CH.

For example, in other embodiments the compounds have one of thefollowing structures (I′i) or (I′j):

wherein A, R¹, R^(2a), R^(2b), R^(2c), R^(3a) and R^(4a) are as definedaccording to any of the foregoing embodiments. In some more specificembodiments of compounds (I′i) or (I′j), R¹ has one of the followingstructures:

In other specific embodiments of compounds (I′i) or (I′j), R^(3a) andR^(4a) are each independently H or C₁-C₆ alkyl, such as methyl. Forexample, in some embodiments R^(3a) and R^(4a) are each H. In otherembodiments, R^(3a) is H and R^(4a) is C₁-C₆ alkyl, such as methyl. Indifferent embodiments, R^(3a) and R^(4a) are each C₁-C₆ alkyl, such asmethyl.

Some embodiments of the compounds include more than one stereoisomer.Other embodiments are directed to a single stereoisomer. In someembodiments the compounds are racemic (e.g., mixture of atropisomers),while in other embodiments the compounds are substantially a singleisomer, for example a substantially purified atropisomer.

In various different embodiments, the compound has one of the structuresset forth in Table 1 below. General methods by which the compounds maybe prepared are described below.

TABLE 1 Representative Compounds No. Structure Name  1

5-(4- acryloylpiperazin- 1-yl)-7-chloro-9- fluoro-8-(2-fluoro-6-hydroxyphenyl)- 3,4- dihydropyrimido [4,5-b]quinolin- 2(1H)-one  2

10-(4- acryloylpiperazin- 1-yl)-8-chloro-6- fluoro-7-(2-fluoro-6-hydroxyphenyl)- 3,4- dihydropyrimido [5,4-b]quinolin- 2(1H)-one  3

10-(4- acryloylpiperazin- 1-yl)-8-chloro-6- fluoro-7-(2-fluoro-6-hydroxyphenyl)- 3,4-dihydrobenzo [b][1,6] naphthyridin- 1(2H)-one  4

5-(4- acryloylpiperazin- 1-yl)-7-chloro-9- fluoro-8-(2-fluoro-6-hydroxyphenyl)- 3,4-dihydrobenzo [b][1,8] naphthyridin- 2(1H)-one  5

10-(4- acryloylpiperazin- 1-yl)-8-chloro-6- fluoro-7-(2-fluoro-6-hydroxyphenyl)- 2H-[1,4]oxazino [3,2-b]quinolin- 3(4H)-one  6

10-(4- acryloylpiperazin- 1-yl)-8-chloro-6- fluoro-7-(2-fluoro-6-hydroxyphenyl)- 3,4- dihydropyrazino [2,3-b]quinolin- 2(1H)-one  7

10-(4- acryloylpiperazin- 1-yl)-8-chloro-6- fluoro-7-(2-fluoro-6-hydroxyphenyl)- 1H-[1,4]oxazino [2,3-b]quinolin- 2(3H)-one  8

10-(4- acryloylpiperazin- 1-yl)-8-chloro-6- fluoro-7-(2-fluoro-6-hydroxyphenyl)- 3,4-dihydrobenzo [b][1,5] naphthyridin- 2(1H)-one  9

1-(4-(8-chloro-6- fluoro-7-(2-fluoro- 6-hydroxyphenyl)- 2,3-dihydro-1H-[1,4]oxazino[2,3- b]quinolin-10-yl) piperazin-1-yl) prop-2-en-1-one 10

1-(4-(8-chloro-6- fluoro-7-(2-fluoro- 6-hydroxyphenyl)- 3,4-dihydro-2H-[1,4]oxazino[3,2- b]quinolin-10-yl) piperazin-1-yl) prop-2-en-1-one 11

1-(4-(8-chloro-6- fluoro-7-(2-fluoro- 6-hydroxyphenyl)- 2,3-dihydro-[1,4]dioxino[2,3- b]quinolin-10-yl) piperazin-1-yl) prop-2-en-1-one 12

10-(4- acryloylpiperazin- 1-yl)-8-chloro-6- fluoro-7-(2- fluoro-6-hydroxyphenyl) benzo[b][1,6] naphthyridin- 1(2H)-one 13

5-(4- acryloylpiperazin- 1-yl)-7-chloro-9- fluoro-8- (2-fluoro-6-hydroxyphenyl) benzo[b][1,8] naphthyridin- 2(1H)-one 14

10-(4- acryloylpiperazin- 1-yl)-8-chloro-6- fluoro-7- (2-fluoro-6-hydroxyphenyl) pyrazino[2,3-b] quinolin-3(4H)- one 15

1-(4-(8-chloro-6- fluoro-7-(2-fluoro- 6-hydroxyphenyl) pyrazino[2,3-b]quinolin-10-yl) piperazin-1-yl) prop-2-en-1-one 16

1-(4-(7-chloro-5- fluoro-6-(2-fluoro- 6-hydroxyphenyl)- 1,2,3,4-tetrahydroacridin- 9-yl)piperazin-1- yl)prop-2-en-1-one 17

1-(4-(8-chloro-6- fluoro-7-(2-fluoro- 6-hydroxyphenyl)-2-methyl-1,2,3,4- tetrahydrobenzo [b][1,6] naphthyridin-10-yl)piperazin-1- yl)prop-2-en-1-one 18

1-(4-(7-chloro-9- fluoro-8-(2-fluoro- 6-hydroxyphenyl)-2-methyl-1,2,3,4- tetrahydrobenzo [b][1,7] naphthyridin-5-yl)piperazin-1- yl)prop-2-en-1-one 19

10-(4- acryloylpiperazin- 1-yl)-8-chloro-6- fluoro-7-(2-fluoro-6-hydroxyphenyl)- 2-methyl-3,4- dihydrobenzo[b] [1,6]naphthyridin-1(2H)-one 20

9-(4- acryloylpiperazin- 1-yl)-7-chloro-5- fluoro-6-(2-fluoro-6-hydroxyphenyl)- 1H-imidazo [4,5-b]quinolin- 2(3H)-one 21

1-(4-(7-chloro-5- fluoro-6-(2-fluoro- 6-hydroxyphenyl) thiazolo[4,5-b]quinolin- 9-yl)piperazin- 1-yl)prop-2-en- 1-one

It is understood that in the present description, combinations ofsubstituents and/or variables of the depicted formulae are permissibleonly if such contributions result in stable compounds.

Furthermore, all compounds of the invention (i.e., compounds ofstructure (I)) which exist in free base or acid form can be converted totheir pharmaceutically acceptable salts by treatment with theappropriate inorganic or organic base or acid by methods known to oneskilled in the art. Salts of the compounds of the invention can beconverted to their free base or acid form by standard techniques.

Methods for making compounds of compounds of structure (I):

or a pharmaceutically acceptable salt, stereoisomer or prodrug thereof,wherein R¹, R^(2a), R^(2b), R^(2c), R^(3a), R^(3b), R^(4a), R^(4b), A,G¹, G², L¹, L², m¹, m² and E are as defined herein, are known in theart. It is understood that one skilled in the art may be able to makethese compounds by methods known to one skilled in the art or bycombinations of known procedures or modification of the same. Ingeneral, starting components may be obtained from sources such as SigmaAldrich, Lancaster Synthesis, Inc., Maybridge, Matrix Scientific, TCI,and Fluorochem USA, etc. or synthesized according to sources known tothose skilled in the art (see, for example, Advanced Organic Chemistry:Reactions, Mechanisms, and Structure, 5th edition (Wiley, December2000)) or prepared as described in this invention.

Embodiments of the compound of structure (I) (e.g., compound A-18) canbe prepared according to General Reaction Scheme 1 (“Method A”), whereinR¹, R^(3a), R^(3b), R^(4a), R^(4b), R⁹ and R¹⁰ are as defined herein, xand y are each independently 0, 1 or 2, and R′ is alkyl. Referring toGeneral Reaction Scheme 1, compounds of structure A-1 are purchased fromcommercial sources and reduced under appropriate conditions to formaniline A-2. A-2 is then treated with 2,2,2-trichloroethane-1,1-diol andsodium sulfate to yield A-3, which is subsequently cyclized by treatmentwith concentrated sulfuric acid. Ring-opening oxidation of A-4 thenyields A-5, which can be optionally chlorinated to yield A-6 when achloro substituent in the R^(2b) position is desired. Esterification ofA-6 yields A-7, which is used as described below.

Separately, A-8 is aminated with an appropriate alkyl amine to yield A-9which is then acylated to form A-10. Cyclization of A-10 by treatmentwith an appropriate base yields A-11, which is subsequentlydecarboxylated to yield A-12. A-12 is then treated with A-7 under acidicconditions to yield A-13. Chlorination of A-13 provides A-14, which isreacted with an appropriate cyclic amine (A-15) to yield A-16. Thedesired R¹ substituent may then be added by way of Suzuki coupling toyield A-17. Removal of the boc protecting group, followed by reactionwith an appropriately substituted acryloyl chloride yields the desiredcompound A-18.

Embodiments of the compound of structure (I) (e.g., compound B-8) can beprepared according to General Reaction Scheme 2 (“Method B”), whereinR¹, R^(3a), R^(3b), R^(4a), R^(4b), R⁹ and R¹⁰ are as defined herein,and x and y are each independently 0, 1 or 2. Referring to GeneralReaction Scheme 2, compounds of structure A-6 are prepared as describedabove. Treatment of A-6 with an appropriate activated carbonate (e.g.,(Cl₃CO)₂CO) provides B-1, which upon treatment with malononitrile yieldsquinolone B-2. Chlorination of B-2 provides B-3, and the heterocyclicmoiety is then installed by treatment with an appropriate heterocycle(e.g., A-15). The desired R¹ substituent may then be added by way ofSuzuki coupling to yield B-5. The desired tricyclic core is thenprovided by reduction of the nitrile group of B-5 to yield B-6, followedby cyclization with a carbodiimide reagent to yield B-7. Removal of theboc protecting group, followed by reaction with an appropriatelysubstituted acryloyl chloride yields the desired compound B-8.

Additional synthetic methods are provided in the Examples. It will beapparent to one of ordinary skill in the art that all compounds ofstructure (I) can be prepared according to one or more of the methodsdescribed herein or otherwise known in the art. It will also be apparentthat in some instances it will be necessary to use a differentlysubstituted starting material and/or protecting groups to arrive at thedesired compound when following the general procedures described herein.Various substituents may also be added at various points in thesynthetic scheme to prepare the desired compound.

Further, one skilled in the art will recognize that certainmodifications to the above schemes and those provided in the examplesare possible to prepare different embodiments of compounds of structure(I). For example, for ease of illustration the General Reaction Schemesabove depict preparation of compounds of structure (I) wherein R^(2a),R^(2b) and R^(2c) are fluoro, chloro and H, respectively. However, itwill be apparent to one of ordinary skill in the art that differentlysubstituted compounds of structure (I) can be prepared according thegeneral methods provided herein by using differently substitutedstarting materials and/or adding the desired substituent using methodsknown in the art.

One of ordinary skill in the art will also readily recognize thatcompounds wherein L¹ is NR⁵ can be prepared by substituting theheterocycle illustrated in the above schemes with a heterocycle havingthe following structure:

where R is H, a protecting group or C₁-C₆alkyl.

It will also be appreciated by those skilled in the art that in theprocesses for preparing the compounds the functional groups ofintermediate compounds may need to be protected by suitable protectinggroups. Such functional groups include, but are not limited to, hydroxy,amino, mercapto and carboxylic acid. Suitable protecting groups forhydroxy include trialkylsilyl or diarylalkylsilyl (for example,t-butyldimethylsilyl, t-butyldiphenylsilyl or trimethylsilyl),tetrahydropyranyl, benzyl, and the like. Suitable protecting groups foramino, amidino and guanidino include t-butoxycarbonyl,benzyloxycarbonyl, and the like. Suitable protecting groups for mercaptoinclude —C(O)—R″ (where R″ is alkyl, aryl or arylalkyl),p-methoxybenzyl, trityl and the like. Suitable protecting groups forcarboxylic acid include alkyl, aryl or arylalkyl esters. Protectinggroups are optionally added or removed in accordance with standardtechniques, which are known to one skilled in the art and as describedherein. The use of protecting groups is described in detail in Green, T.W. and P. G. M. Wutz, Protective Groups in Organic Synthesis (1999), 3rdEd., Wiley. As one of skill in the art would appreciate, the protectinggroup may also be a polymer resin such as a Wang resin, Rink resin or a2-chlorotrityl-chloride resin.

It will also be appreciated by those skilled in the art, although suchprotected derivatives of compounds of this invention may not possesspharmacological activity as such, they may be administered to a mammaland thereafter metabolized in the body to form compounds of theinvention which are pharmacologically active. Such derivatives maytherefore be described as “prodrugs”. All prodrugs of compounds of thisinvention are included within the scope of the invention.

Pharmaceutical Compositions

Other embodiments are directed to pharmaceutical compositions. Thepharmaceutical composition comprises any one (or more) of the foregoingcompounds and a pharmaceutically acceptable carrier. In someembodiments, the pharmaceutical composition is formulated for oraladministration. In other embodiments, the pharmaceutical composition isformulated for injection. In still more embodiments, the pharmaceuticalcompositions comprise a compound as disclosed herein and an additionaltherapeutic agent (e.g., anticancer agent). Non-limiting examples ofsuch therapeutic agents are described herein below.

Suitable routes of administration include, but are not limited to, oral,intravenous, rectal, aerosol, parenteral, ophthalmic, pulmonary,transmucosal, transdermal, vaginal, otic, nasal, and topicaladministration. In addition, by way of example only, parenteral deliveryincludes intramuscular, subcutaneous, intravenous, intramedullaryinjections, as well as intrathecal, direct intraventricular,intraperitoneal, intralymphatic, and intranasal injections.

In certain embodiments, a compound as described herein is administeredin a local rather than systemic manner, for example, via injection ofthe compound directly into an organ, often in a depot preparation orsustained release formulation. In specific embodiments, long actingformulations are administered by implantation (for examplesubcutaneously or intramuscularly) or by intramuscular injection.Furthermore, in other embodiments, the drug is delivered in a targeteddrug delivery system, for example, in a liposome coated withorgan-specific antibody. In such embodiments, the liposomes are targetedto and taken up selectively by the organ. In yet other embodiments, thecompound as described herein is provided in the form of a rapid releaseformulation, in the form of an extended release formulation, or in theform of an intermediate release formulation. In yet other embodiments,the compound described herein is administered topically.

The compounds according to the invention are effective over a widedosage range. For example, in the treatment of adult humans, dosagesfrom 0.01 to 1000 mg, from 0.5 to 100 mg, from 1 to 50 mg per day, andfrom 5 to 40 mg per day are examples of dosages that are used in someembodiments. An exemplary dosage is 10 to 30 mg per day. The exactdosage will depend upon the route of administration, the form in whichthe compound is administered, the subject to be treated, the body weightof the subject to be treated, and the preference and experience of theattending physician.

In some embodiments, a compound of the invention is administered in asingle dose. Typically, such administration will be by injection, e.g.,intravenous injection, in order to introduce the agent quickly. However,other routes are used as appropriate. A single dose of a compound of theinvention may also be used for treatment of an acute condition.

In some embodiments, a compound of the invention is administered inmultiple doses. In some embodiments, dosing is about once, twice, threetimes, four times, five times, six times, or more than six times perday. In other embodiments, dosing is about once a month, once every twoweeks, once a week, or once every other day. In another embodiment acompound of the invention and another agent are administered togetherabout once per day to about 6 times per day. In another embodiment theadministration of a compound of the invention and an agent continues forless than about 7 days. In yet another embodiment the administrationcontinues for more than about 6, 10, 14, 28 days, two months, sixmonths, or one year. In some cases, continuous dosing is achieved andmaintained as long as necessary.

Administration of the compounds of the invention may continue as long asnecessary. In some embodiments, a compound of the invention isadministered for more than 1, 2, 3, 4, 5, 6, 7, 14, or 28 days. In someembodiments, a compound of the invention is administered for less than28, 14, 7, 6, 5, 4, 3, 2, or 1 day. In some embodiments, a compound ofthe invention is administered chronically on an ongoing basis, e.g., forthe treatment of chronic effects.

In some embodiments, the compounds of the invention are administered indosages. It is known in the art that due to intersubject variability incompound pharmacokinetics, individualization of dosing regimen isnecessary for optimal therapy. Dosing for a compound of the inventionmay be found by routine experimentation in light of the instantdisclosure.

In some embodiments, the compounds described herein are formulated intopharmaceutical compositions. In specific embodiments, pharmaceuticalcompositions are formulated in a conventional manner using one or morephysiologically acceptable carriers comprising excipients andauxiliaries which facilitate processing of the active compounds intopreparations which can be used pharmaceutically. Proper formulation isdependent upon the route of administration chosen. Any pharmaceuticallyacceptable techniques, carriers, and excipients are used as suitable toformulate the pharmaceutical compositions described herein: Remington:The Science and Practice of Pharmacy, Nineteenth Ed (Easton, Pa.: MackPublishing Company, 1995); Hoover, John E., Remington's PharmaceuticalSciences, Mack Publishing Co., Easton, Pa. 1975; Liberman, H. A. andLachman, L., Eds., Pharmaceutical Dosage Forms, Marcel Decker, New York,N.Y., 1980; and Pharmaceutical Dosage Forms and Drug Delivery Systems,Seventh Ed. (Lippincott Williams & Wilkins 1999).

Provided herein are pharmaceutical compositions comprising a compound ofstructure (I) and a pharmaceutically acceptable diluent(s),excipient(s), or carrier(s). In certain embodiments, the compoundsdescribed are administered as pharmaceutical compositions in whichcompounds of structure (I) are mixed with other active ingredients, asin combination therapy. Encompassed herein are all combinations ofactives set forth in the combination therapies section below andthroughout this disclosure. In specific embodiments, the pharmaceuticalcompositions include one or more compounds of structure (I).

A pharmaceutical composition, as used herein, refers to a mixture of acompound of structure (I) with other chemical components, such ascarriers, stabilizers, diluents, dispersing agents, suspending agents,thickening agents, and/or excipients. In certain embodiments, thepharmaceutical composition facilitates administration of the compound toan organism. In some embodiments, practicing the methods of treatment oruse provided herein, therapeutically effective amounts of compounds ofstructure (I) provided herein are administered in a pharmaceuticalcomposition to a mammal having a disease, disorder or medical conditionto be treated. In specific embodiments, the mammal is a human. Incertain embodiments, therapeutically effective amounts vary depending onthe severity of the disease, the age and relative health of the subject,the potency of the compound used and other factors. The compoundsdescribed herein are used singly or in combination with one or moretherapeutic agents as components of mixtures.

In one embodiment, one or more compounds of structure (I) is formulatedin an aqueous solutions. In specific embodiments, the aqueous solutionis selected from, by way of example only, a physiologically compatiblebuffer, such as Hank's solution, Ringer's solution, or physiologicalsaline buffer. In other embodiments, one or more compound of structure(I) is/are formulated for transmucosal administration. In specificembodiments, transmucosal formulations include penetrants that areappropriate to the barrier to be permeated. In still other embodimentswherein the compounds described herein are formulated for otherparenteral injections, appropriate formulations include aqueous ornonaqueous solutions. In specific embodiments, such solutions includephysiologically compatible buffers and/or excipients.

In another embodiment, compounds described herein are formulated fororal administration. Compounds described herein are formulated bycombining the active compounds (compounds of structure (I)) with, e.g.,pharmaceutically acceptable carriers or excipients. In variousembodiments, the compounds described herein are formulated in oraldosage forms that include, by way of example only, tablets, powders,pills, dragees, capsules, liquids, gels, syrups, elixirs, slurries,suspensions and the like.

In certain embodiments, pharmaceutical preparations for oral use areobtained by mixing one or more solid excipient with one or more of thecompounds described herein, optionally grinding the resulting mixture,and processing the mixture of granules, after adding suitableauxiliaries, if desired, to obtain tablets or dragee cores. Suitableexcipients are, in particular, fillers such as sugars, includinglactose, sucrose, mannitol, or sorbitol; cellulose preparations such as:for example, maize starch, wheat starch, rice starch, potato starch,gelatin, gum tragacanth, methylcellulose, microcrystalline cellulose,hydroxypropylmethylcellulose, sodium carboxymethylcellulose; or otherssuch as: polyvinylpyrrolidone (PVP or povidone) or calcium phosphate. Inspecific embodiments, disintegrating agents are optionally added.Disintegrating agents include, by way of example only, cross-linkedcroscarmellose sodium, polyvinylpyrrolidone, agar, or alginic acid or asalt thereof such as sodium alginate.

In one embodiment, dosage forms, such as dragee cores and tablets, areprovided with one or more suitable coating. In specific embodiments,concentrated sugar solutions are used for coating the dosage form. Thesugar solutions, optionally contain additional components, such as byway of example only, gum arabic, talc, polyvinylpyrrolidone, carbopolgel, polyethylene glycol, and/or titanium dioxide, lacquer solutions,and suitable organic solvents or solvent mixtures. Dyestuffs and/orpigments are also optionally added to the coatings for identificationpurposes. Additionally, the dyestuffs and/or pigments are optionallyutilized to characterize different combinations of active compounddoses.

In certain embodiments, therapeutically effective amounts of at leastone of the compounds described herein are formulated into other oraldosage forms. Oral dosage forms include push-fit capsules made ofgelatin, as well as soft, sealed capsules made of gelatin and aplasticizer, such as glycerol or sorbitol. In specific embodiments,push-fit capsules contain the active ingredients in admixture with oneor more filler. Fillers include, by way of example only, lactose,binders such as starches, and/or lubricants such as talc or magnesiumstearate and, optionally, stabilizers. In other embodiments, softcapsules, contain one or more active compound that is dissolved orsuspended in a suitable liquid. Suitable liquids include, by way ofexample only, one or more fatty oil, liquid paraffin, or liquidpolyethylene glycol. In addition, stabilizers are optionally added.

In other embodiments, therapeutically effective amounts of at least oneof the compounds described herein are formulated for buccal orsublingual administration. Formulations suitable for buccal orsublingual administration include, by way of example only, tablets,lozenges, or gels. In still other embodiments, the compounds describedherein are formulated for parental injection, including formulationssuitable for bolus injection or continuous infusion. In specificembodiments, formulations for injection are presented in unit dosageform (e.g., in ampoules) or in multi-dose containers. Preservatives are,optionally, added to the injection formulations. In still otherembodiments, the pharmaceutical compositions are formulated in a formsuitable for parenteral injection as sterile suspensions, solutions oremulsions in oily or aqueous vehicles. Parenteral injection formulationsoptionally contain formulatory agents such as suspending, stabilizingand/or dispersing agents. In specific embodiments, pharmaceuticalformulations for parenteral administration include aqueous solutions ofthe active compounds in water-soluble form. In additional embodiments,suspensions of the active compounds (e.g., compounds of structure (I))are prepared as appropriate oily injection suspensions. Suitablelipophilic solvents or vehicles for use in the pharmaceuticalcompositions described herein include, by way of example only, fattyoils such as sesame oil, or synthetic fatty acid esters, such as ethyloleate or triglycerides, or liposomes. In certain specific embodiments,aqueous injection suspensions contain substances which increase theviscosity of the suspension, such as sodium carboxymethyl cellulose,sorbitol, or dextran. Optionally, the suspension contains suitablestabilizers or agents which increase the solubility of the compounds toallow for the preparation of highly concentrated solutions.Alternatively, in other embodiments, the active ingredient is in powderform for constitution with a suitable vehicle, e.g., sterilepyrogen-free water, before use.

In still other embodiments, the compounds of structure (I) areadministered topically. The compounds described herein are formulatedinto a variety of topically administrable compositions, such assolutions, suspensions, lotions, gels, pastes, medicated sticks, balms,creams or ointments. Such pharmaceutical compositions optionally containsolubilizers, stabilizers, tonicity enhancing agents, buffers andpreservatives.

In yet other embodiments, the compounds of structure (I) are formulatedfor transdermal administration. In specific embodiments, transdermalformulations employ transdermal delivery devices and transdermaldelivery patches and can be lipophilic emulsions or buffered, aqueoussolutions, dissolved and/or dispersed in a polymer or an adhesive. Invarious embodiments, such patches are constructed for continuous,pulsatile, or on demand delivery of pharmaceutical agents. In additionalembodiments, the transdermal delivery of the compounds of structure (I)is accomplished by means of iontophoretic patches and the like. Incertain embodiments, transdermal patches provide controlled delivery ofthe compounds of structure (I). In specific embodiments, the rate ofabsorption is slowed by using rate-controlling membranes or by trappingthe compound within a polymer matrix or gel. In alternative embodiments,absorption enhancers are used to increase absorption. Absorptionenhancers or carriers include absorbable pharmaceutically acceptablesolvents that assist passage through the skin. For example, in oneembodiment, transdermal devices are in the form of a bandage comprisinga backing member, a reservoir containing the compound optionally withcarriers, optionally a rate controlling barrier to deliver the compoundto the skin of the host at a controlled and predetermined rate over aprolonged period of time, and means to secure the device to the skin.

In other embodiments, the compounds of structure (I) are formulated foradministration by inhalation. Various forms suitable for administrationby inhalation include, but are not limited to, aerosols, mists orpowders. Pharmaceutical compositions of any of compound of structure (I)are conveniently delivered in the form of an aerosol spray presentationfrom pressurized packs or a nebuliser, with the use of a suitablepropellant (e.g., dichlorodifluoromethane, trichlorofluoromethane,dichlorotetrafluoroethane, carbon dioxide or other suitable gas). Inspecific embodiments, the dosage unit of a pressurized aerosol isdetermined by providing a valve to deliver a metered amount. In certainembodiments, capsules and cartridges of, such as, by way of exampleonly, gelatin for use in an inhaler or insufflator are formulatedcontaining a powder mix of the compound and a suitable powder base suchas lactose or starch.

In still other embodiments, the compounds of structure (I) areformulated in rectal compositions such as enemas, rectal gels, rectalfoams, rectal aerosols, suppositories, jelly suppositories, or retentionenemas, containing conventional suppository bases such as cocoa butteror other glycerides, as well as synthetic polymers such aspolyvinylpyrrolidone, PEG, and the like. In suppository forms of thecompositions, a low-melting wax such as, but not limited to, a mixtureof fatty acid glycerides, optionally in combination with cocoa butter isfirst melted.

In certain embodiments, pharmaceutical compositions are formulated inany conventional manner using one or more physiologically acceptablecarriers comprising excipients and auxiliaries which facilitateprocessing of the active compounds into preparations which can be usedpharmaceutically. Proper formulation is dependent upon the route ofadministration chosen. Any pharmaceutically acceptable techniques,carriers, and excipients are optionally used as suitable. Pharmaceuticalcompositions comprising a compound of structure (I) are manufactured ina conventional manner, such as, by way of example only, by means ofconventional mixing, dissolving, granulating, dragee-making, levigating,emulsifying, encapsulating, entrapping or compression processes.

Pharmaceutical compositions include at least one pharmaceuticallyacceptable carrier, diluent or excipient and at least one compound ofstructure (I), described herein as an active ingredient. The activeingredient is in free-acid or free-base form, or in a pharmaceuticallyacceptable salt form. In addition, the methods and pharmaceuticalcompositions described herein include the use of N-oxides, crystallineforms (also known as polymorphs), as well as active metabolites of thesecompounds having the same type of activity. All tautomers of thecompounds described herein are included within the scope of thecompounds presented herein. Additionally, the compounds described hereinencompass unsolvated as well as solvated forms with pharmaceuticallyacceptable solvents such as water, ethanol, and the like. The solvatedforms of the compounds presented herein are also considered to bedisclosed herein. In addition, the pharmaceutical compositionsoptionally include other medicinal or pharmaceutical agents, carriers,adjuvants, such as preserving, stabilizing, wetting or emulsifyingagents, solution promoters, salts for regulating the osmotic pressure,buffers, and/or other therapeutically valuable substances.

Methods for the preparation of compositions comprising the compoundsdescribed herein include formulating the compounds with one or moreinert, pharmaceutically acceptable excipients or carriers to form asolid, semi-solid or liquid. Solid compositions include, but are notlimited to, powders, tablets, dispersible granules, capsules, cachets,and suppositories. Liquid compositions include solutions in which acompound is dissolved, emulsions comprising a compound, or a solutioncontaining liposomes, micelles, or nanoparticles comprising a compoundas disclosed herein. Semi-solid compositions include, but are notlimited to, gels, suspensions and creams. The form of the pharmaceuticalcompositions described herein include liquid solutions or suspensions,solid forms suitable for solution or suspension in a liquid prior touse, or as emulsions. These compositions also optionally contain minoramounts of nontoxic, auxiliary substances, such as wetting oremulsifying agents, pH buffering agents, and so forth.

In some embodiments, pharmaceutical composition comprising at least onecompound of structure (I) illustratively takes the form of a liquidwhere the agents are present in solution, in suspension or both.Typically when the composition is administered as a solution orsuspension a first portion of the agent is present in solution and asecond portion of the agent is present in particulate form, insuspension in a liquid matrix. In some embodiments, a liquid compositionincludes a gel formulation. In other embodiments, the liquid compositionis aqueous.

In certain embodiments, useful aqueous suspensions contain one or morepolymers as suspending agents. Useful polymers include water-solublepolymers such as cellulosic polymers, e.g., hydroxypropylmethylcellulose, and water-insoluble polymers such as cross-linkedcarboxyl-containing polymers. Certain pharmaceutical compositionsdescribed herein comprise a mucoadhesive polymer, selected for examplefrom carboxymethylcellulose, carbomer (acrylic acid polymer),poly(methylmethacrylate), polyacrylamide, polycarbophil, acrylicacid/butyl acrylate copolymer, sodium alginate and dextran.

Useful pharmaceutical compositions also, optionally, includesolubilizing agents to aid in the solubility of a compound of structure(I). The term “solubilizing agent” generally includes agents that resultin formation of a micellar solution or a true solution of the agent.Certain acceptable nonionic surfactants, for example polysorbate 80, areuseful as solubilizing agents, as can ophthalmically acceptable glycols,polyglycols, e.g., polyethylene glycol 400, and glycol ethers.

Furthermore, useful pharmaceutical compositions optionally include oneor more pH adjusting agents or buffering agents, including acids such asacetic, boric, citric, lactic, phosphoric and hydrochloric acids; basessuch as sodium hydroxide, sodium phosphate, sodium borate, sodiumcitrate, sodium acetate, sodium lactate andtris-hydroxymethylaminomethane; and buffers such as citrate/dextrose,sodium bicarbonate and ammonium chloride. Such acids, bases and buffersare included in an amount required to maintain pH of the composition inan acceptable range.

Additionally, useful compositions also, optionally, include one or moresalts in an amount required to bring osmolality of the composition intoan acceptable range. Such salts include those having sodium, potassiumor ammonium cations and chloride, citrate, ascorbate, borate, phosphate,bicarbonate, sulfate, thiosulfate or bisulfite anions; suitable saltsinclude sodium chloride, potassium chloride, sodium thiosulfate, sodiumbisulfite and ammonium sulfate.

Other useful pharmaceutical compositions optionally include one or morepreservatives to inhibit microbial activity. Suitable preservativesinclude mercury-containing substances such as merfen and thiomersal;stabilized chlorine dioxide; and quaternary ammonium compounds such asbenzalkonium chloride, cetyltrimethylammonium bromide andcetylpyridinium chloride.

Still other useful compositions include one or more surfactants toenhance physical stability or for other purposes. Suitable nonionicsurfactants include polyoxyethylene fatty acid glycerides and vegetableoils, e.g., polyoxyethylene (60) hydrogenated castor oil; andpolyoxyethylene alkylethers and alkylphenyl ethers, e.g., octoxynol 10,octoxynol 40.

Still other useful compositions include one or more antioxidants toenhance chemical stability where required. Suitable antioxidantsinclude, by way of example only, ascorbic acid and sodium metabisulfite.

In certain embodiments, aqueous suspension compositions are packaged insingle-dose non-reclosable containers. Alternatively, multiple-dosereclosable containers are used, in which case it is typical to include apreservative in the composition.

In alternative embodiments, other delivery systems for hydrophobicpharmaceutical compounds are employed. Liposomes and emulsions areexamples of delivery vehicles or carriers useful herein. In certainembodiments, organic solvents such as N-methylpyrrolidone are alsoemployed. In additional embodiments, the compounds described herein aredelivered using a sustained-release system, such as semipermeablematrices of solid hydrophobic polymers containing the therapeutic agent.Various sustained-release materials are useful herein. In someembodiments, sustained-release capsules release the compounds for a fewweeks up to over 100 days. Depending on the chemical nature and thebiological stability of the therapeutic reagent, additional strategiesfor protein stabilization are employed.

In certain embodiments, the formulations described herein comprise oneor more antioxidants, metal chelating agents, thiol containing compoundsand/or other general stabilizing agents. Examples of such stabilizingagents, include, but are not limited to: (a) about 0.5% to about 2% w/vglycerol, (b) about 0.1% to about 1% w/v methionine, (c) about 0.1% toabout 2% w/v monothioglycerol, (d) about 1 mM to about 10 mM EDTA, (e)about 0.01% to about 2% w/v ascorbic acid, (f) 0.003% to about 0.02% w/vpolysorbate 80, (g) 0.001% to about 0.05% w/v. polysorbate 20, (h)arginine, (i) heparin, (j) dextran sulfate, (k) cyclodextrins, (l)pentosan polysulfate and other heparinoids, (m) divalent cations such asmagnesium and zinc; or (n) combinations thereof.

In some embodiments, the concentration of one or more compounds providedin the pharmaceutical compositions of the present invention is less than100%, 90%, 80%, 70%, 60%, 50%, 40%, 30%, 20%, 19%, 18%, 17%, 16%, 15%,14%, 13%, 12%, 11%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, 0.5%, 0.4%,0.3%, 0.2%, 0.1%, 0.09%, 0.08%, 0.07%, 0.06%, 0.05%, 0.04%, 0.03%,0.02%, 0.01%, 0.009%, 0.008%, 0.007%, 0.006%, 0.005%, 0.004%, 0.003%,0.002%, 0.001%, 0.0009%, 0.0008%, 0.0007%, 0.0006%, 0.0005%, 0.0004%,0.0003%, 0.0002%, or 0.0001% w/w, w/v or v/v.

In some embodiments, the concentration of one or more compounds of theinvention is greater than 90%, 80%, 70%, 60%, 50%, 40%, 30%, 20%,19.75%, 19.50%, 19.25% 19%, 18.75%, 18.50%, 18.25% 18%, 17.75%, 17.50%,17.25% 17%, 16.75%, 16.50%, 16.25% 16%, 15.75%, 15.50%, 15.25% 15%,14.75%, 14.50%, 14.25% 14%, 13.75%, 13.50%, 13.25% 13%, 12.75%, 12.50%,12.25% 12%, 11.75%, 11.50%, 11.25% 11%, 10.75%, 10.50%, 10.25% 10%,9.75%, 9.50%, 9.25% 9%, 8.75%, 8.50%, 8.25% 8%, 7.75%, 7.50%, 7.25% 7%,6.75%, 6.50%, 6.25% 6%, 5.75%, 5.50%, 5.25% 5%, 4.75%, 4.50%, 4.25%, 4%,3.75%, 3.50%, 3.25%, 3%, 2.75%, 2.50%, 2.25%, 2%, 1.75%, 1.50%, 125%,1%, 0.5%, 0.4%, 0.3%, 0.2%, 0.1%, 0.09%, 0.08%, 0.07%, 0.06%, 0.05%,0.04%, 0.03%, 0.02%, 0.01%, 0.009%, 0.008%, 0.007%, 0.006%, 0.005%,0.004%, 0.003%, 0.002%, 0.001%, 0.0009%, 0.0008%, 0.0007%, 0.0006%,0.0005%, 0.0004%, 0.0003%, 0.0002%, or 0.0001% w/w, w/v, or v/v.

In some embodiments, the concentration of one or more compounds of theinvention is in the range from approximately 0.0001% to approximately50%, approximately 0.001% to approximately 40%, approximately 0.01% toapproximately 30%, approximately 0.02% to approximately 29%,approximately 0.03% to approximately 28%, approximately 0.04% toapproximately 27%, approximately 0.05% to approximately 26%,approximately 0.06% to approximately 25%, approximately 0.07% toapproximately 24%, approximately 0.08% to approximately 23%,approximately 0.09% to approximately 22%, approximately 0.1% toapproximately 21%, approximately 0.2% to approximately 20%,approximately 0.3% to approximately 19%, approximately 0.4% toapproximately 18%, approximately 0.5% to approximately 17%,approximately 0.6% to approximately 16%, approximately 0.7% toapproximately 15%, approximately 0.8% to approximately 14%,approximately 0.9% to approximately 12%, approximately 1% toapproximately 10% w/w, w/v or v/v.

In some embodiments, the concentration of one or more compounds of theinvention is in the range from approximately 0.001% to approximately10%, approximately 0.01% to approximately 5%, approximately 0.02% toapproximately 4.5%, approximately 0.03% to approximately 4%,approximately 0.04% to approximately 3.5%, approximately 0.05% toapproximately 3%, approximately 0.06% to approximately 2.5%,approximately 0.07% to approximately 2%, approximately 0.08% toapproximately 1.5%, approximately 0.09% to approximately 1%,approximately 0.1% to approximately 0.9% w/w, w/v or v/v.

In some embodiments, the amount of one or more compounds of theinvention is equal to or less than 10 g, 9.5 g, 9.0 g, 8.5 g, 8.0 g, 7.5g, 7.0 g, 6.5 g, 6.0 g, 5.5 g, 5.0 g, 4.5 g, 4.0 g, 3.5 g, 3.0 g, 2.5 g,2.0 g, 1.5 g, 1.0 g, 0.95 g, 0.9 g, 0.85 g, 0.8 g, 0.75 g, 0.7 g, 0.65g, 0.6 g, 0.55 g, 0.5 g, 0.45 g, 0.4 g, 0.35 g, 0.3 g, 0.25 g, 0.2 g,0.15 g, 0.1 g, 0.09 g, 0.08 g, 0.07 g, 0.06 g, 0.05 g, 0.04 g, 0.03 g,0.02 g, 0.01 g, 0.009 g, 0.008 g, 0.007 g, 0.006 g, 0.005 g, 0.004 g,0.003 g, 0.002 g, 0.001 g, 0.0009 g, 0.0008 g, 0.0007 g, 0.0006 g,0.0005 g, 0.0004 g, 0.0003 g, 0.0002 g, or 0.0001 g.

In some embodiments, the amount of one or more compounds of theinvention is more than 0.0001 g, 0.0002 g, 0.0003 g, 0.0004 g, 0.0005 g,0.0006 g, 0.0007 g, 0.0008 g, 0.0009 g, 0.001 g, 0.0015 g, 0.002 g,0.0025 g, 0.003 g, 0.0035 g, 0.004 g, 0.0045 g, 0.005 g, 0.0055 g, 0.006g, 0.0065 g, 0.007 g, 0.0075 g, 0.008 g, 0.0085 g, 0.009 g, 0.0095 g,0.01 g, 0.015 g, 0.02 g, 0.025 g, 0.03 g, 0.035 g, 0.04 g, 0.045 g, 0.05g, 0.055 g, 0.06 g, 0.065 g, 0.07 g, 0.075 g, 0.08 g, 0.085 g, 0.09 g,0.095 g, 0.1 g, 0.15 g, 0.2 g, 0.25 g, 0.3 g, 0.35 g, 0.4 g, 0.45 g, 0.5g, 0.55 g, 0.6 g, 0.65 g, 0.7 g, 0.75 g, 0.8 g, 0.85 g, 0.9 g, 0.95 g, 1g, 1.5 g, 2 g, 2.5, 3 g, 3.5, 4 g, 4.5 g, 5 g, 5.5 g, 6 g, 6.5 g, 7 g,7.5 g, 8 g, 8.5 g, 9 g, 9.5 g, or 10 g.

In some embodiments, the amount of one or more compounds of theinvention is in the range of 0.0001-10 g, 0.0005-9 g, 0.001-8 g, 0.005-7g, 0.01-6 g, 0.05-5 g, 0.1-4 g, 0.5-4 g, or 1-3 g.

Kits/Articles of Manufacture

For use in the therapeutic applications described herein, kits andarticles of manufacture are also provided. In some embodiments, suchkits comprise a carrier, package, or container that is compartmentalizedto receive one or more containers such as vials, tubes, and the like,each of the container(s) comprising one of the separate elements to beused in a method described herein. Suitable containers include, forexample, bottles, vials, syringes, and test tubes. The containers areformed from a variety of materials such as glass or plastic.

The articles of manufacture provided herein contain packaging materials.Packaging materials for use in packaging pharmaceutical products includethose found in, e.g., U.S. Pat. Nos. 5,323,907, 5,052,558 and 5,033,252.Examples of pharmaceutical packaging materials include, but are notlimited to, blister packs, bottles, tubes, inhalers, pumps, bags, vials,containers, syringes, bottles, and any packaging material suitable for aselected formulation and intended mode of administration and treatment.For example, the container(s) includes one or more compounds describedherein, optionally in a composition or in combination with another agentas disclosed herein. The container(s) optionally have a sterile accessport (for example the container is an intravenous solution bag or a vialhaving a stopper pierceable by a hypodermic injection needle). Such kitsoptionally comprising a compound with an identifying description orlabel or instructions relating to its use in the methods describedherein.

For example, a kit typically includes one or more additional containers,each with one or more of various materials (such as reagents, optionallyin concentrated form, and/or devices) desirable from a commercial anduser standpoint for use of a compound described herein. Non-limitingexamples of such materials include, but not limited to, buffers,diluents, filters, needles, syringes; carrier, package, container, vialand/or tube labels listing contents and/or instructions for use, andpackage inserts with instructions for use. A set of instructions willalso typically be included. A label is optionally on or associated withthe container. For example, a label is on a container when letters,numbers or other characters forming the label are attached, molded oretched into the container itself, a label is associated with a containerwhen it is present within a receptacle or carrier that also holds thecontainer, e.g., as a package insert. In addition, a label is used toindicate that the contents are to be used for a specific therapeuticapplication. In addition, the label indicates directions for use of thecontents, such as in the methods described herein. In certainembodiments, the pharmaceutical compositions is presented in a pack ordispenser device which contains one or more unit dosage forms containinga compound provided herein. The pack for example contains metal orplastic foil, such as a blister pack. Or, the pack or dispenser deviceis accompanied by instructions for administration. Or, the pack ordispenser is accompanied with a notice associated with the container inform prescribed by a governmental agency regulating the manufacture,use, or sale of pharmaceuticals, which notice is reflective of approvalby the agency of the form of the drug for human or veterinaryadministration. Such notice, for example, is the labeling approved bythe U.S. Food and Drug Administration for prescription drugs, or theapproved product insert. In some embodiments, compositions containing acompound provided herein formulated in a compatible pharmaceuticalcarrier are prepared, placed in an appropriate container, and labeledfor treatment of an indicated condition.

Methods

Embodiments of the present invention provide a method of inhibitingRAS-mediated cell signaling comprising contacting a cell with aneffective amount of one or more compounds disclosed herein. Inhibitionof RAS-mediated signal transduction can be assessed and demonstrated bya wide variety of ways known in the art. Non-limiting examples include ashowing of (a) a decrease in GTPase activity of RAS; (b) a decrease inGTP binding affinity or an increase in GDP binding affinity; (c) anincrease in K off of GTP or a decrease in K off of GDP; (d) a decreasein the levels of signaling transduction molecules downstream in the RASpathway, such as a decrease in pMEK level; and/or (e) a decrease inbinding of RAS complex to downstream signaling molecules including butnot limited to Raf. Kits and commercially available assays can beutilized for determining one or more of the above.

Embodiments also provide methods of using the compounds orpharmaceutical compositions of the present invention to treat diseaseconditions, including but not limited to conditions implicated by G12CKRAS, HRAS or NRAS mutation, G12C HRAS mutation and/or G12C NRASmutation (e.g., cancer).

In some embodiments, a method for treatment of cancer is provided, themethod comprising administering an effective amount of any of theforegoing pharmaceutical compositions comprising a compound of structure(I) to a subject in need thereof. In some embodiments, the cancer ismediated by a KRAS, HRAS or NRAS G12C mutation. In other embodiments,the cancer is pancreatic cancer, colon cancer, MYH associated polyposis,colorectal cancer or lung cancer.

In some embodiments the invention provides method of treating a disorderin a subject in need thereof, wherein the said method comprisesdetermining if the subject has a KRAS, HRAS or NRAS G12C mutation and ifthe subject is determined to have the KRAS, HRAS or NRAS G12C mutation,then administering to the subject a therapeutically effective dose of atleast one compound of structure (I) or a pharmaceutically acceptablesalt, ester, prodrug, tautomer, solvate, hydrate or derivative thereof.

Embodiments of the disclosed compounds strongly inhibitanchorage-independent cell growth and therefore have the potential toinhibit tumor metastasis. Accordingly, in another embodiment thedisclosure provides a method for inhibiting tumor metastasis, the methodcomprising administering an effective amount a pharmaceuticalcomposition of comprising any of the compounds disclosed herein and apharmaceutically acceptable carrier to a subject in need thereof.

KRAS, HRAS or NRAS G12C mutations have also been identified inhematological malignancies (e.g., cancers that affect blood, bone marrowand/or lymph nodes). Accordingly, certain embodiments are directed toadministration of a disclosed compounds (e.g., in the form of apharmaceutical composition) to a patient in need of treatment of ahematological malignancy. Such malignancies include, but are not limitedto leukemias and lymphomas. For example, the presently disclosedcompounds can be used for treatment of diseases such as Acutelymphoblastic leukemia (ALL), Acute myelogenous leukemia (AML), Chroniclymphocytic leukemia (CLL), small lymphocytic lymphoma (SLL), Chronicmyelogenous leukemia (CML), Acute monocytic leukemia (AMoL) and/or otherleukemias. In other embodiments, the compounds are useful for treatmentof lymphomas such as all subtypes of Hodgkin's lymphoma or non-Hodgkin'slymphoma.

Determining whether a tumor or cancer comprises a G12C KRAS, HRAS orNRAS mutation can be undertaken by assessing the nucleotide sequenceencoding the KRAS, HRAS or NRAS protein, by assessing the amino acidsequence of the KRAS, HRAS or NRAS protein, or by assessing thecharacteristics of a putative KRAS, HRAS or NRAS mutant protein. Thesequence of wild-type human KRAS, HRAS or NRAS is known in the art,(e.g. Accession No. NP203524).

Methods for detecting a mutation in a KRAS, HRAS or NRAS nucleotidesequence are known by those of skill in the art. These methods include,but are not limited to, polymeRASe chain reaction-restriction fragmentlength polymorphism (PCR-RFLP) assays, polymeRASe chain reaction-singlestrand conformation polymorphism (PCR-SSCP) assays, real-time PCRassays, PCR sequencing, mutant allele-specific PCR amplification (MASA)assays, direct sequencing, primer extension reactions, electrophoresis,oligonucleotide ligation assays, hybridization assays, TaqMan assays,SNP genotyping assays, high resolution melting assays and microarrayanalyses. In some embodiments, samples are evaluated for G12C KRAS, HRASor NRAS mutations by real-time PCR. In real-time PCR, fluorescent probesspecific for the KRAS, HRAS or NRAS G12C mutation are used. When amutation is present, the probe binds and fluorescence is detected. Insome embodiments, the KRAS, HRAS or NRAS G12C mutation is identifiedusing a direct sequencing method of specific regions (e.g., exon 2and/or exon 3) in the KRAS, HRAS or NRAS gene. This technique willidentify all possible mutations in the region sequenced.

Methods for detecting a mutation in a KRAS, HRAS or NRAS protein areknown by those of skill in the art. These methods include, but are notlimited to, detection of a KRAS, HRAS or NRAS mutant using a bindingagent (e.g., an antibody) specific for the mutant protein, proteinelectrophoresis and Western blotting, and direct peptide sequencing.

Methods for determining whether a tumor or cancer comprises a G12C KRAS,HRAS or NRAS mutation can use a variety of samples. In some embodiments,the sample is taken from a subject having a tumor or cancer. In someembodiments, the sample is taken from a subject having a cancer ortumor. In some embodiments, the sample is a fresh tumor/cancer sample.In some embodiments, the sample is a frozen tumor/cancer sample. In someembodiments, the sample is a formalin-fixed paraffin-embedded sample. Insome embodiments, the sample is processed to a cell lysate. In someembodiments, the sample is processed to DNA or RNA.

Embodiments also relate to a method of treating a hyperproliferativedisorder in a mammal that comprises administering to said mammal atherapeutically effective amount of a compound of the present invention,or a pharmaceutically acceptable salt, ester, prodrug, solvate, hydrateor derivative thereof. In some embodiments, said method relates to thetreatment of cancer such as acute myeloid leukemia, cancer inadolescents, adrenocortical carcinoma childhood, AIDS-related cancers(e.g. Lymphoma and Kaposi's Sarcoma), anal cancer, appendix cancer,astrocytomas, atypical teratoid, basal cell carcinoma, bile duct cancer,bladder cancer, bone cancer, brain stem glioma, brain tumor, breastcancer, bronchial tumors, burkitt lymphoma, carcinoid tumor, atypicalteratoid, embryonal tumors, germ cell tumor, primary lymphoma, cervicalcancer, childhood cancers, chordoma, cardiac tumors, chronic lymphocyticleukemia (CLL), chronic myelogenous leukemia (CML), chronicmyleoproliferative disorders, colon cancer, colorectal cancer,craniopharyngioma, cutaneous T-cell lymphoma, extrahepatic ductalcarcinoma in situ (DCIS), embryonal tumors, CNS cancer, endometrialcancer, ependymoma, esophageal cancer, esthesioneuroblastoma, ewingsarcoma, extracranial germ cell tumor, extragonadal germ cell tumor, eyecancer, fibrous histiocytoma of bone, gall bladder cancer, gastriccancer, gastrointestinal carcinoid tumor, gastrointestinal stromaltumors (GIST), germ cell tumor, gestational trophoblastic tumor, hairycell leukemia, head and neck cancer, heart cancer, liver cancer,Hodgkin's lymphoma, hypopharyngeal cancer, intraocular melanoma, isletcell tumors, pancreatic neuroendocrine tumors, kidney cancer, laryngealcancer, lip and oral cavity cancer, liver cancer, lobular carcinoma insitu (LCIS), lung cancer, lymphoma, metastatic squamous neck cancer withoccult primary, midline tract carcinoma, mouth cancer multiple endocrineneoplasia syndromes, multiple myeloma/plasma cell neoplasm, mycosisfungoides, myelodysplastic syndromes, myelodysplastic/myeloproliferativeneoplasms, multiple myeloma, merkel cell carcinoma, malignantmesothelioma, malignant fibrous histiocytoma of bone and osteosarcoma,nasal cavity and paranasal sinus cancer, nasopharyngeal cancer,neuroblastoma, non-Hodgkin's lymphoma, non-small cell lung cancer(NSCLC), oral cancer, lip and oral cavity cancer, oropharyngeal cancer,ovarian cancer, pancreatic cancer, papillomatosis, paraganglioma,paranasal sinus and nasal cavity cancer, parathyroid cancer, penilecancer, pharyngeal cancer, pleuropulmonary blastoma, primary centralnervous system (CNS) lymphoma, prostate cancer, rectal cancer,transitional cell cancer, retinoblastoma, rhabdomyosarcoma, salivarygland cancer, skin cancer, stomach (gastric) cancer, small cell lungcancer, small intestine cancer, soft tissue sarcoma, T-Cell lymphoma,testicular cancer, throat cancer, thymoma and thymic carcinoma, thyroidcancer, transitional cell cancer of the renal pelvis and ureter,trophoblastic tumor, unusual cancers of childhood, urethral cancer,uterine sarcoma, vaginal cancer, vulvar cancer, or Viral-Induced cancer.In some embodiments, said method relates to the treatment of anon-cancerous hyperproliferative disorder such as benign hyperplasia ofthe skin (e.g., psoriasis), restenosis, or prostate (e.g., benignprostatic hypertrophy (BPH)).

In certain particular embodiments, the invention relates to methods fortreatment of lung cancers, the methods comprise administering aneffective amount of any of the above described compound (or apharmaceutical composition comprising the same) to a subject in needthereof. In certain embodiments the lung cancer is a non-small cell lungcarcinoma (NSCLC), for example adenocarcinoma, squamous-cell lungcarcinoma or large-cell lung carcinoma. In other embodiments, the lungcancer is a small cell lung carcinoma. Other lung cancers treatable withthe disclosed compounds include, but are not limited to, glandulartumors, carcinoid tumors and undifferentiated carcinomas.

Subjects that can be treated with compounds of the invention, orpharmaceutically acceptable salt, ester, prodrug, solvate, tautomer,hydrate or derivative of said compounds, according to the methods ofthis invention include, for example, subjects that have been diagnosedas having acute myeloid leukemia, acute myeloid leukemia, cancer inadolescents, adrenocortical carcinoma childhood, AIDS-related cancers(e.g. Lymphoma and Kaposi's Sarcoma), anal cancer, appendix cancer,astrocytomas, atypical teratoid, basal cell carcinoma, bile duct cancer,bladder cancer, bone cancer, brain stem glioma, brain tumor, breastcancer, bronchial tumors, burkitt lymphoma, carcinoid tumor, atypicalteratoid, embryonal tumors, germ cell tumor, primary lymphoma, cervicalcancer, childhood cancers, chordoma, cardiac tumors, chronic lymphocyticleukemia (CLL), chronic myelogenous leukemia (CML), chronicmyleoproliferative disorders, colon cancer, colorectal cancer,craniopharyngioma, cutaneous T-cell lymphoma, extrahepatic ductalcarcinoma in situ (DCIS), embryonal tumors, CNS cancer, endometrialcancer, ependymoma, esophageal cancer, esthesioneuroblastoma, ewingsarcoma, extracranial germ cell tumor, extragonadal germ cell tumor, eyecancer, fibrous histiocytoma of bone, gall bladder cancer, gastriccancer, gastrointestinal carcinoid tumor, gastrointestinal stromaltumors (GIST), germ cell tumor, gestational trophoblastic tumor, hairycell leukemia, head and neck cancer, heart cancer, liver cancer,Hodgkin's lymphoma, hypopharyngeal cancer, intraocular melanoma, isletcell tumors, pancreatic neuroendocrine tumors, kidney cancer, laryngealcancer, lip and oral cavity cancer, liver cancer, lobular carcinoma insitu (LCIS), lung cancer, lymphoma, metastatic squamous neck cancer withoccult primary, midline tract carcinoma, mouth cancer multiple endocrineneoplasia syndromes, multiple myeloma/plasma cell neoplasm, mycosisfungoides, myelodysplastic syndromes, myelodysplastic/myeloproliferativeneoplasms, multiple myeloma, merkel cell carcinoma, malignantmesothelioma, malignant fibrous histiocytoma of bone and osteosarcoma,nasal cavity and paranasal sinus cancer, nasopharyngeal cancer,neuroblastoma, non-Hodgkin's lymphoma, non-small cell lung cancer(NSCLC), oral cancer, lip and oral cavity cancer, oropharyngeal cancer,ovarian cancer, pancreatic cancer, papillomatosis, paraganglioma,paranasal sinus and nasal cavity cancer, parathyroid cancer, penilecancer, pharyngeal cancer, pleuropulmonary blastoma, primary centralnervous system (CNS) lymphoma, prostate cancer, rectal cancer,transitional cell cancer, retinoblastoma, rhabdomyosarcoma, salivarygland cancer, skin cancer, stomach (gastric) cancer, small cell lungcancer, small intestine cancer, soft tissue sarcoma, T-Cell lymphoma,testicular cancer, throat cancer, thymoma and thymic carcinoma, thyroidcancer, transitional cell cancer of the renal pelvis and ureter,trophoblastic tumor, unusual cancers of childhood, urethral cancer,uterine sarcoma, vaginal cancer, vulvar cancer, or Viral-Induced cancer.In some embodiments subjects that are treated with the compounds of theinvention include subjects that have been diagnosed as having anon-cancerous hyperproliferative disorder such as benign hyperplasia ofthe skin (e.g., psoriasis), restenosis, or prostate (e.g., benignprostatic hypertrophy (BPH)).

Embodiments further provide methods of modulating a G12C Mutant KRAS,HRAS or NRAS protein activity by contacting the protein with aneffective amount of a compound of the invention. Modulation can beinhibiting or activating protein activity. In some embodiments, theinvention provides methods of inhibiting protein activity by contactingthe G12C Mutant KRAS, HRAS or NRAS protein with an effective amount of acompound of the invention in solution. In some embodiments, theinvention provides methods of inhibiting the G12C Mutant KRAS, HRAS orNRAS protein activity by contacting a cell, tissue, organ that expressthe protein of interest. In some embodiments, the invention providesmethods of inhibiting protein activity in subject including but notlimited to rodents and mammal (e.g., human) by administering into thesubject an effective amount of a compound of the invention. In someembodiments, the percentage modulation exceeds 25%, 30%, 40%, 50%, 60%,70%, 80%, or 90%. In some embodiments, the percentage of inhibitingexceeds 25%, 30%, 40%, 50%, 60%, 70%, 80%, or 90%.

In some embodiments, the invention provides methods of inhibiting KRAS,HRAS or NRAS G12C activity in a cell by contacting said cell with anamount of a compound of the invention sufficient to inhibit the activityof KRAS, HRAS or NRAS G12C in said cell. In some embodiments, theinvention provides methods of inhibiting KRAS, HRAS or NRAS G12Cactivity in a tissue by contacting said tissue with an amount of acompound of the invention sufficient to inhibit the activity of KRAS,HRAS or NRAS G12C in said tissue. In some embodiments, the inventionprovides methods of inhibiting KRAS, HRAS or NRAS G12C activity in anorganism by contacting said organism with an amount of a compound of theinvention sufficient to inhibit the activity of KRAS, HRAS or NRAS G12Cin said organism. In some embodiments, the invention provides methods ofinhibiting KRAS, HRAS or NRAS G12C activity in an animal by contactingsaid animal with an amount of a compound of the invention sufficient toinhibit the activity of KRAS, HRAS or NRAS G12C in said animal. In someembodiments, the invention provides methods of inhibiting KRAS, HRAS orNRAS G12C activity in a mammal by contacting said mammal with an amountof a compound of the invention sufficient to inhibit the activity ofKRAS, HRAS or NRAS G12C in said mammal. In some embodiments, theinvention provides methods of inhibiting KRAS, HRAS or NRAS G12Cactivity in a human by contacting said human with an amount of acompound of the invention sufficient to inhibit the activity of KRAS,HRAS or NRAS G12C in said human. In other embodiments, the presentinvention provides methods of treating a disease mediated by KRAS, HRASor NRAS G12C activity in a subject in need of such treatment.

In different embodiments, the present invention also provides methodsfor combination therapies in which an agent known to modulate otherpathways, or other components of the same pathway, or even overlappingsets of target enzymes are used in combination with a compound of thepresent invention, or a pharmaceutically acceptable salt, ester,prodrug, solvate, tautomer, hydrate or derivative thereof. In oneaspect, such therapy includes but is not limited to the combination ofone or more compounds of the invention with chemotherapeutic agents,therapeutic antibodies, and radiation treatment, to provide asynergistic or additive therapeutic effect.

Many chemotherapeutics are presently known in the art and can be used incombination with the compounds of the invention. In some embodiments,the chemotherapeutic is selected from the group consisting of mitoticinhibitors, alkylating agents, anti-metabolites, intercalatingantibiotics, growth factor inhibitors, cell cycle inhibitors, enzymes,topoisomeRASe inhibitors, biological response modifiers, anti-hormones,angiogenesis inhibitors, and anti-androgens.

Non-limiting examples are chemotherapeutic agents, cytotoxic agents, andnon-peptide small molecules such as Gleevec® (Imatinib Mesylate),Velcade® (bortezomib), Casodex (bicalutamide), Iressa® (gefitinib), andAdriamycin as well as a host of chemotherapeutic agents. Non-limitingexamples of chemotherapeutic agents include alkylating agents such asthiotepa and cyclosphosphamide (CYTOXAN™); alkyl sulfonates such asbusulfan, improsulfan and piposulfan; aziridines such as benzodopa,carboquone, meturedopa, and uredopa; ethylenimines and methylamelaminesincluding altretamine, triethylenemelamine, trietylenephosphoramide,triethylenethiophosphaoramide and trimethylolomelamine; nitrogenmustards such as chlorambucil, chlornaphazine, cholophosphamide,estramustine, ifosfamide, mechlorethamine, mechlorethamine oxidehydrochloride, melphalan, novembichin, phenesterine, prednimustine,trofosfamide, uracil mustard; nitrosureas such as carmustine,chlorozotocin, fotemustine, lomustine, nimustine, ranimustine;antibiotics such as aclacinomysins, actinomycin, authramycin, azaserine,bleomycins, cactinomycin, calicheamicin, carabicin, carminomycin,carzinophilin, Casodex™, chromomycins, dactinomycin, daunorubicin,detorubicin, 6-diazo-5-oxo-L-norleucine, doxorubicin, epirubicin,esorubicin, idarubicin, marcellomycin, mitomycins, mycophenolic acid,nogalamycin, olivomycins, peplomycin, potfiromycin, puromycin,quelamycin, rodorubicin, streptonigrin, streptozocin, tubercidin,ubenimex, zinostatin, zorubicin; anti-metabolites such as methotrexateand 5-fluorouracil (5-FU); folic acid analogues such as denopterin,methotrexate, pteropterin, trimetrexate; purine analogs such asfludarabine, 6-mercaptopurine, thiamiprine, thioguanine; pyrimidineanalogs such as ancitabine, azacitidine, 6-azauridine, carmofur,cytarabine, dideoxyuridine, doxifluridine, enocitabine, floxuridine,androgens such as calusterone, dromostanolone propionate, epitiostanol,mepitiostane, testolactone; anti-adrenals such as aminoglutethimide,mitotane, trilostane; folic acid replenisher such as frolinic acid;aceglatone; aldophosphamide glycoside; aminolevulinic acid; amsacrine;bestrabucil; bisantrene; edatraxate; defofamine; demecolcine;diaziquone; elfomithine; elliptinium acetate; etoglucid; galliumnitrate; hydroxyurea; lentinan; lonidamine; mitoguazone; mitoxantrone;mopidamol; nitracrine; pentostatin; phenamet; pirarubicin; podophyllinicacid; 2-ethylhydrazide; procarbazine; PSK®; razoxane; sizofiran;spirogermanium; tenuazonic acid; triaziquone;2,2′,2″-trichlorotriethylamine; urethan; vindesine; dacarbazine;mannomustine; mitobronitol; mitolactol; pipobroman; gacytosine;arabinoside (“Ara-C”); cyclophosphamide; thiotepa; taxanes, e.g.paclitaxel (TAXOL™, Bristol-Myers Squibb Oncology, Princeton, N.J.) anddocetaxel (TAXOTERE™, Rhone-Poulenc Rorer, Antony, France); retinoicacid; esperamicins; capecitabine; and pharmaceutically acceptable salts,acids or derivatives of any of the above. Also included as suitablechemotherapeutic cell conditioners are anti-hormonal agents that act toregulate or inhibit hormone action on tumors such as anti-estrogensincluding for example tamoxifen, (Nolvadex™), raloxifene, aromataseinhibiting 4(5)-imidazoles, 4-hydroxytamoxifen, trioxifene, keoxifene,LY 117018, onapristone, and toremifene (Fareston); and anti-androgenssuch as flutamide, nilutamide, bicalutamide, leuprolide, and goserelin;chlorambucil; gemcitabine; 6-thioguanine; mercaptopurine; methotrexate;platinum analogs such as cisplatin and carboplatin; vinblastine;platinum; etoposide (VP-16); ifosfamide; mitomycin C; mitoxantrone;vincristine; vinorelbine; navelbine; novantrone; teniposide; daunomycin;aminopterin; xeloda; ibandronate; camptothecin-11 (CPT-11);topoisomeRASe inhibitor RFS 2000; difluoromethylornithine (DMFO). Wheredesired, the compounds or pharmaceutical composition of the presentinvention can be used in combination with commonly prescribedanti-cancer drugs such as Herceptin®, Avastin®, Erbitux®, Rituxan®,Taxol®, Arimidex®, Taxotere®, ABVD, AVICINE, Abagovomab, Acridinecarboxamide, Adecatumumab, 17-N-Allylamino-17-demethoxygeldanamycin,Alpharadin, Alvocidib, 3-Aminopyridine-2-carboxaldehydethiosemicarbazone, Amonafide, Anthracenedione, Anti-CD22 immunotoxins,Antineoplastic, Antitumorigenic herbs, Apaziquone, Atiprimod,Azathioprine, Belotecan, Bendamustine, BIBW 2992, Biricodar,Brostallicin, Bryostatin, Buthionine sulfoximine, CBV (chemotherapy),Calyculin, cell-cycle nonspecific antineoplastic agents, Dichloroaceticacid, Discodermolide, Elsamitrucin, Enocitabine, Epothilone, Eribulin,Everolimus, Exatecan, Exisulind, Ferruginol, Forodesine, Fosfestrol, ICEchemotherapy regimen, IT-101, Imexon, Imiquimod, Indolocarbazole,Irofulven, Laniquidar, Larotaxel, Lenalidomide, Lucanthone, Lurtotecan,Mafosfamide, Mitozolomide, Nafoxidine, Nedaplatin, Olaparib, Ortataxel,PAC-1, Pawpaw, Pixantrone, Proteasome inhibitor, Rebeccamycin,Resiquimod, Rubitecan, SN-38, Salinosporamide A, Sapacitabine, StanfordV, Swainsonine, Talaporfin, Tariquidar, Tegafur-uracil, Temodar,Tesetaxel, Triplatin tetranitrate, Tris(2-chloroethyl)amine,Troxacitabine, Uramustine, Vadimezan, Vinflunine, ZD6126 or Zosuquidar.

Embodiments further relate to a method for using the compounds orpharmaceutical compositions provided herein, in combination withradiation therapy for inhibiting abnormal cell growth or treating thehyperproliferative disorder in the mammal. Techniques for administeringradiation therapy are known in the art, and these techniques can be usedin the combination therapy described herein. The administration of thecompound of the invention in this combination therapy can be determinedas described herein.

Radiation therapy can be administered through one of several methods, ora combination of methods, including without limitation external-beamtherapy, internal radiation therapy, implant radiation, stereotacticradiosurgery, systemic radiation therapy, radiotherapy and permanent ortemporary interstitial brachytherapy. The term “brachytherapy,” as usedherein, refers to radiation therapy delivered by a spatially confinedradioactive material inserted into the body at or near a tumor or otherproliferative tissue disease site. The term is intended withoutlimitation to include exposure to radioactive isotopes (e.g. At-211,I-131, I-125, Y-90, Re-186, Re-188, Sm-153, Bi-212, P-32, andradioactive isotopes of Lu). Suitable radiation sources for use as acell conditioner of the present invention include both solids andliquids. By way of non-limiting example, the radiation source can be aradionuclide, such as I-125, I-131, Yb-169, Ir-192 as a solid source,I-125 as a solid source, or other radionuclides that emit photons, betaparticles, gamma radiation, or other therapeutic rays. The radioactivematerial can also be a fluid made from any solution of radionuclide(s),e.g., a solution of I-125 or I-131, or a radioactive fluid can beproduced using a slurry of a suitable fluid containing small particlesof solid radionuclides, such as Au-198, Y-90. Moreover, theradionuclide(s) can be embodied in a gel or radioactive micro spheres.

Without being limited by any theory, the compounds of the presentinvention can render abnormal cells more sensitive to treatment withradiation for purposes of killing and/or inhibiting the growth of suchcells. Accordingly, this invention further relates to a method forsensitizing abnormal cells in a mammal to treatment with radiation whichcomprises administering to the mammal an amount of a compound of thepresent invention or pharmaceutically acceptable salt, ester, prodrug,solvate, hydrate or derivative thereof, which amount is effective issensitizing abnormal cells to treatment with radiation. The amount ofthe compound, salt, or solvate in this method can be determinedaccording to the means for ascertaining effective amounts of suchcompounds described herein.

The compounds or pharmaceutical compositions of the invention can beused in combination with an amount of one or more substances selectedfrom anti-angiogenesis agents, signal transduction inhibitors,antiproliferative agents, glycolysis inhibitors, or autophagyinhibitors.

Anti-angiogenesis agents, such as MMP-2 (matrix-metalloproteinase 2)inhibitors, MMP-9 (matrix-metalloprotienase 9) inhibitors, and COX-11(cyclooxygenase 11) inhibitors, can be used in conjunction with acompound of the invention and pharmaceutical compositions describedherein. Anti-angiogenesis agents include, for example, rapamycin,temsirolimus (CCI-779), everolimus (RAD001), sorafenib, sunitinib, andbevacizumab. Examples of useful COX-II inhibitors include CELEBREX™(alecoxib), valdecoxib, and rofecoxib. Examples of useful matrixmetalloproteinase inhibitors are described in WO 96/33172 (publishedOct. 24, 1996), WO 96/27583 (published Mar. 7, 1996), European PatentApplication No. 97304971.1 (filed Jul. 8, 1997), European PatentApplication No. 99308617.2 (filed Oct. 29, 1999), WO 98/07697 (publishedFeb. 26, 1998), WO 98/03516 (published Jan. 29, 1998), WO 98/34918(published Aug. 13, 1998), WO 98/34915 (published Aug. 13, 1998), WO98/33768 (published Aug. 6, 1998), WO 98/30566 (published Jul. 16,1998), European Patent Publication 606,046 (published Jul. 13, 1994),European Patent Publication 931, 788 (published Jul. 28, 1999), WO90/05719 (published May 31, 1990), WO 99/52910 (published Oct. 21,1999), WO 99/52889 (published Oct. 21, 1999), WO 99/29667 (publishedJun. 17, 1999), PCT International Application No. PCT/IB98/01113 (filedJul. 21, 1998), European Patent Application No. 99302232.1 (filed Mar.25, 1999), Great Britain Patent Application No. 9912961.1 (filed Jun. 3,1999), U.S. Provisional Application No. 60/148,464 (filed Aug. 12,1999), U.S. Pat. No. 5,863,949 (issued Jan. 26, 1999), U.S. Pat. No.5,861,510 (issued Jan. 19, 1999), and European Patent Publication780,386 (published Jun. 25, 1997), all of which are incorporated hereinin their entireties by reference. Preferred MMP-2 and MMP-9 inhibitorsare those that have little or no activity inhibiting MMP-1. Morepreferred, are those that selectively inhibit MMP-2 and/or AMP-9relative to the other matrix-metalloproteinases (i.e., MAP-1, MMP-3,MMP-4, MMP-5, MMP-6, MMP-7, MMP-8, MMP-10, MMP-11, MMP-12, and MMP-13).Some specific examples of MMP inhibitors useful in the invention areAG-3340, RO 32-3555, and RS 13-0830.

Autophagy inhibitors include, but are not limited to chloroquine,3-methyladenine, hydroxychloroquine (Plaquenil™), bafilomycin A1,5-amino-4-imidazole carboxamide riboside (AICAR), okadaic acid,autophagy-suppressive algal toxins which inhibit protein phosphatases oftype 2A or type 1, analogues of cAMP, and drugs which elevate cAMPlevels such as adenosine, LY204002, N6-mercaptopurine riboside, andvinblastine. In addition, antisense or siRNA that inhibits expression ofproteins including but not limited to ATG5 (which are implicated inautophagy), may also be used.

Different embodiments relate to a method of and to a pharmaceuticalcomposition for treating a cardiovascular disease in a mammal whichcomprises an amount of a compound of the invention, or apharmaceutically acceptable salt, ester, prodrug, solvate, tautomer,hydrate or derivative thereof, or an isotopically-labeled derivativethereof, and an amount of one or more therapeutic agents use for thetreatment of cardiovascular diseases.

Exemplary agents for use in cardiovascular disease applications areanti-thrombotic agents, e.g., prostacyclin and salicylates, thrombolyticagents, e.g., streptokinase, urokinase, tissue plasminogen activator(TPA) and anisoylated plasminogen-streptokinase activator complex(APSAC), anti-platelets agents, e.g., acetyl-salicylic acid (ASA) andclopidrogel, vasodilating agents, e.g., nitrates, calcium channelblocking drugs, anti-proliferative agents, e.g., colchicine andalkylating agents, intercalating agents, growth modulating factors suchas interleukins, transformation growth factor-beta and congeners ofplatelet derived growth factor, monoclonal antibodies directed againstgrowth factors, anti-inflammatory agents, both steroidal andnon-steroidal, and other agents that can modulate vessel tone, function,arteriosclerosis, and the healing response to vessel or organ injurypost intervention. Antibiotics can also be included in combinations orcoatings comprised by the invention. Moreover, a coating can be used toeffect therapeutic delivery focally within the vessel wall. Byincorporation of the active agent in a swellable polymer, the activeagent will be released upon swelling of the polymer.

In some embodiments, the compounds described herein are formulated oradministered in conjunction with liquid or solid tissue barriers alsoknown as lubricants. Examples of tissue barriers include, but are notlimited to, polysaccharides, polyglycans, seprafilm, interceed andhyaluronic acid.

In some embodiments, medicaments which are administered in conjunctionwith the compounds described herein include any suitable drugs usefullydelivered by inhalation for example, analgesics, e.g. codeine,dihydromorphine, ergotamine, fentanyl or morphine; anginal preparations,e.g. diltiazem; antiallergics, e.g. cromoglycate, ketotifen ornedocromil; anti-infectives, e.g. cephalosporins, penicillins,streptomycin, sulphonamides, tetracyclines or pentamidine;antihistamines, e.g. methapyrilene; anti-inflammatories, e.g.beclomethasone, flunisolide, budesonide, tipredane, triamcinoloneacetonide or fluticasone; antitussives, e.g. noscapine; bronchodilators,e.g. ephedrine, adrenaline, fenoterol, formoterol, isoprenaline,metaproterenol, phenylephrine, phenylpropanolamine, pirbuterol,reproterol, rimiterol, salbutamol, salmeterol, terbutalin, isoetharine,tulobuterol, orciprenaline or(−)-4-amino-3,5-dichloro-α-[[[6-[2-(2-pyridinyl)ethoxy]hexyl]-amino]methyl]benzenemethanol;diuretics, e.g. amiloride; anticholinergics e.g. ipratropium, atropineor oxitropium; hormones, e.g. cortisone, hydrocortisone or prednisolone;xanthines e.g. aminophylline, choline theophyllinate, lysinetheophyllinate or theophylline; and therapeutic proteins and peptides,e.g. insulin or glucagon. It will be clear to a person skilled in theart that, where appropriate, the medicaments are used in the form ofsalts (e.g. as alkali metal or amine salts or as acid addition salts) oras esters (e.g. lower alkyl esters) or as solvates (e.g. hydrates) tooptimize the activity and/or stability of the medicament.

Other exemplary therapeutic agents useful for a combination therapyinclude but are not limited to agents as described above, radiationtherapy, hormone antagonists, hormones and their releasing factors,thyroid and antithyroid drugs, estrogens and progestins, androgens,adrenocorticotropic hormone; adrenocortical steroids and their syntheticanalogs; inhibitors of the synthesis and actions of adrenocorticalhormones, insulin, oral hypoglycemic agents, and the pharmacology of theendocrine pancreas, agents affecting calcification and bone turnover:calcium, phosphate, parathyroid hormone, vitamin D, calcitonin, vitaminssuch as water-soluble vitamins, vitamin B complex, ascorbic acid,fat-soluble vitamins, vitamins A, K, and E, growth factors, cytokines,chemokines, muscarinic receptor agonists and antagonists;anticholinesteRASe agents; agents acting at the neuromuscular junctionand/or autonomic ganglia; catecholamines, sympathomimetic drugs, andadrenergic receptor agonists or antagonists; and 5-hydroxytryptamine(5-HT, serotonin) receptor agonists and antagonists.

Therapeutic agents can also include agents for pain and inflammationsuch as histamine and histamine antagonists, bradykinin and bradykininantagonists, 5-hydroxytryptamine (serotonin), lipid substances that aregenerated by biotransformation of the products of the selectivehydrolysis of membrane phospholipids, eicosanoids, prostaglandins,thromboxanes, leukotrienes, aspirin, nonsteroidal anti-inflammatoryagents, analgesic-antipyretic agents, agents that inhibit the synthesisof prostaglandins and thromboxanes, selective inhibitors of theinducible cyclooxygenase, selective inhibitors of the induciblecyclooxygenase-2, autacoids, paracrine hormones, somatostatin, gastrin,cytokines that mediate interactions involved in humoral and cellularimmune responses, lipid-derived autacoids, eicosanoids, β-adrenergicagonists, ipratropium, glucocorticoids, methylxanthines, sodium channelblockers, opioid receptor agonists, calcium channel blockers, membranestabilizers and leukotriene inhibitors.

Additional therapeutic agents contemplated herein include diuretics,vasopressin, agents affecting the renal conservation of water, rennin,angiotensin, agents useful in the treatment of myocardial ischemia,anti-hypertensive agents, angiotensin converting enzyme inhibitors,β-adrenergic receptor antagonists, agents for the treatment ofhypercholesterolemia, and agents for the treatment of dyslipidemia.

Other therapeutic agents contemplated include drugs used for control ofgastric acidity, agents for the treatment of peptic ulcers, agents forthe treatment of gastroesophageal reflux disease, prokinetic agents,antiemetics, agents used in irritable bowel syndrome, agents used fordiarrhea, agents used for constipation, agents used for inflammatorybowel disease, agents used for biliary disease, agents used forpancreatic disease. Therapeutic agents used to treat protozoaninfections, drugs used to treat Malaria, Amebiasis, Giardiasis,Trichomoniasis, Trypanosomiasis, and/or Leishmaniasis, and/or drugs usedin the chemotherapy of helminthiasis. Other therapeutic agents includeantimicrobial agents, sulfonamides, trimethoprim-sulfamethoxazolequinolones, and agents for urinary tract infections, penicillins,cephalosporins, and other, β-lactam antibiotics, an agent comprising anaminoglycoside, protein synthesis inhibitors, drugs used in thechemotherapy of tuberculosis, mycobacterium avium complex disease, andleprosy, antifungal agents, antiviral agents including nonretroviralagents and antiretroviral agents.

Examples of therapeutic antibodies that can be combined with a compoundof the invention include but are not limited to anti-receptor tyrosinekinase antibodies (cetuximab, panitumumab, tRAStuzumab), anti CD20antibodies (rituximab, tositumomab), and other antibodies such asalemtuzumab, bevacizumab, and gemtuzumab.

Moreover, therapeutic agents used for immunomodulation, such asimmunomodulators, immunosuppressive agents, tolerogens, andimmunostimulants are contemplated by the methods herein. In addition,therapeutic agents acting on the blood and the blood-forming organs,hematopoietic agents, growth factors, minerals, and vitamins,anticoagulant, thrombolytic, and antiplatelet drugs.

For treating renal carcinoma, one may combine a compound of the presentinvention with sorafenib and/or avastin. For treating an endometrialdisorder, one may combine a compound of the present invention withdoxorubincin, taxotere (taxol), and/or cisplatin (carboplatin). Fortreating ovarian cancer, one may combine a compound of the presentinvention with cisplatin (carboplatin), taxotere, doxorubincin,topotecan, and/or tamoxifen. For treating breast cancer, one may combinea compound of the present invention with taxotere (taxol), gemcitabine(capecitabine), tamoxifen, letrozole, tarceva, lapatinib, PD0325901,avastin, herceptin, OSI-906, and/or OSI-930. For treating lung cancer,one may combine a compound of the present invention with taxotere(taxol), gemcitabine, cisplatin, pemetrexed, Tarceva, PD0325901, and/oravastin.

In other embodiments, agents useful in methods for combination therapywith one or more compounds of structure (I) include, but are not limitedto: Erlotinib, Afatinib, Iressa, GDC0941, MLN1117, BYL719 (Alpelisib),BKM120 (Buparlisib), CYT387, GLPG0634, Baricitinib, Lestaurtinib,momelotinib, Pacritinib, Ruxolitinib, TG101348, Crizotinib, tivantinib,AMG337, cabozantinib, foretinib, onartuzumab, NVP-AEW541, Dasatinib,Ponatinib, saracatinib, bosutinib, trametinib, selumetinib, cobimetinib,PD0325901, RO5126766, Axitinib, Bevacizumab, Bostutinib, Cetuximab,Crizotinib, Fostamatinib, Gefitinib, Imatinib, Lapatinib, Lenvatinib,Ibrutinib, Nilotinib, Panitumumab, Pazopanib, Pegaptanib, Ranibizumab,Ruxolitinib, Sorafenib, Sunitinib, SU6656, Trastuzumab, Tofacitinib,Vandetanib, Vemurafenib, Irinotecan, Taxol, Docetaxel, Rapamycin orMLN0128.

Further therapeutic agents that can be combined with a compound of theinvention are found in Goodman and Gilman's “The Pharmacological Basisof Therapeutics” Tenth Edition edited by Hardman, Limbird and Gilman orthe Physician's Desk Reference, both of which are incorporated herein byreference in their entirety.

The compounds described herein can be used in combination with theagents disclosed herein or other suitable agents, depending on thecondition being treated. Hence, in some embodiments the one or morecompounds of the invention will be co-administered with other agents asdescribed above. When used in combination therapy, the compoundsdescribed herein are administered with the second agent simultaneouslyor separately. This administration in combination can includesimultaneous administration of the two agents in the same dosage form,simultaneous administration in separate dosage forms, and separateadministration. That is, a compound described herein and any of theagents described above can be formulated together in the same dosageform and administered simultaneously. Alternatively, a compound of theinvention and any of the agents described above can be simultaneouslyadministered, wherein both the agents are present in separateformulations. In another alternative, a compound of the presentinvention can be administered just followed by and any of the agentsdescribed above, or vice versa. In some embodiments of the separateadministration protocol, a compound of the invention and any of theagents described above are administered a few minutes apart, or a fewhours apart, or a few days apart.

The examples and preparations provided below further illustrate andexemplify the compounds of the present invention and methods ofpreparing such compounds. It is to be understood that the scope of thepresent invention is not limited in any way by the scope of thefollowing examples and preparations. In the following examples, andthroughout the specification and claims, molecules with a single chiralcenter, unless otherwise noted, exist as a racemic mixture. Thosemolecules with two or more chiral centers, unless otherwise noted, existas a racemic mixture of diastereomers. Single enantiomers/diastereomersmay be obtained by methods known to those skilled in the art.

EXAMPLES Example 1 Synthesis of10-(4-acryloylpiperazin-1-yl)-8-chloro-6-fluoro-7-(2-fluoro-6-hydroxyphenyl)-2-methyl-3,4-dihydrobenzo[b][1,6]naphthyridin-1(2h)-one

Example 1 provides an exemplary preparation according to Method A.

3-Bromo-2-fluorobenzenamine

To a mixture of 3-bromo-2-fluorobenzenamine (20.00 g, 90.91 mmol, 1.00eq), EtOH (220.00 mL), AcOH (38.38 g, 639.10 mmol, 36.55 mL, 7.03 eq)and H₂O (88.00 mL), was added Fe (13.20 g, 236.37 mmol, 2.60 eq)portion-wise. The mixture was stirred at 25° C. for 16 h. The reactionmixture was filtered. The filtrate was adjusted pH to 7˜8 with aq.NaOH(2M), and concentrated under reduced pressure. The residue was extractedwith ethyl acetate (50 mL×3). The combined organic layer was washed withbrine (50 mL), dried over Na₂SO₄, filtered and concentrated. The residuewas purified by column chromatography on silica gel (PE/EA=15/1 to 10/1)to afford the desired product (15.12 g, 79.57 mmol, 87.53% yield). ¹HNMR (400 MHz, CHLOROFORM-d): δ 6.91-6.87 (m, 1H), 6.83-6.78 (m, 1H),6.72-6.68 (t, 1H).

N-(3-Bromo-2-fluorophenyl)-2-(hydroxyimino)acetamide

To a mixture of 2,2,2-trichloroethane-1,1-diol (10.45 g, 63.16 mmol,8.23 mL, 1.20 eq) and Na₂SO₄ (67.28 g, 473.67 mmol, 48.06 mL, 9.00 eq)in H₂O (166.00 mL) at 60° C., was added 3-bromo-2-fluorobenzenamine(10.00 g, 52.63 mmol, 1.00 eq). The mixture was stirred at 60° C. for 1h, and then con.HCl (8.80 mL) was added to the mixture. The resultingmixture was stirred at 60° C. for 1 h. NH₂OH.HCl (18.29 g, 263.15 mmol,5.00 eq) was added to this mixture. The resulting mixture was stirred at60° C. for 4 h and then stirred at 100° C. for 16 h. The mixture wasallowed to cool to 25° C., The yellow precipitate was formed. The solidwas collected by filtration, rinsed with water (50 mL×3) and dried toafford the desired product (8.00 g, 30.65 mmol, 58.23% yield) as ayellow solid. ¹H NMR (400 MHz, DMSO-d₆): δ 12.36 (s, 1H), 9.98 (s, 1H),7.83-7.79 (m, 1H), 7.52-7.49 (m, 1H), 7.18-7.14 (m, 1H).

6-Bromo-7-fluoroindoline-2,3-dione

To the conc. H₂SO₄ (130.00 mL) at 60° C., was addedN-(3-bromo-2-fluorophenyl)-2-(hydroxyimino)acetamide (17.00 g, 65.12mmol, 1.00 eq). The mixture was stirred at 90° C. for 1 h. The mixturewas cooled to 25° C. and poured into ice-water (80 mL). The mixture wasextracted with ethyl acetate (100 mL×3). The organic layer was washedwith water (50 mL), saturated NaHCO₃ (50 mL) and brine (50 mL), driedover Na₂SO₄ and concentrated in vacuo. The residue was purified bycolumn chromatography on silica gel (PE/EA=10:1˜5:1) to afford thedesired product (6.00 g, 24.59 mmol, 37.76% yield) as yellow solid. ¹HNMR (400 MHz, DMSO-d₆): δ 11.73 (s, 1H), 7.39-7.36 (m, 1H), 7.31-7.29(m, 1H).

2-Amino-4-bromo-3-fluorobenzoic Acid

To a mixture of 6-bromo-7-fluoroindoline-2,3-dione (6.00 g, 24.59 mmol,1.00 eq) in NaOH (2 M, 111.02 mL, 9.03 eq) at 0° C., was added H₂O₂(17.70 g, 156.13 mmol, 15.00 mL, 30%, 6.35 eq). The mixture was stirredat 25° C. for 16 h. The mixture was quenched with Na₂SO₃ (7 g), and thenthe mixture was acidified with conc.HCl to adjust pH to 2. Theprecipitate was collected by filtration and dried to afford the desiredproduct (5.00 g, 21.37 mmol, 86.89% yield) as a light yellow solid. ¹HNMR (400 MHz, DMSO-d₆): δ 7.48-7.46 (m, 1H), 6.79-6.76 (m, 1H).

2-Amino-4-bromo-5-chloro-3-fluorobenzoic Acid

To a solution of 2-amino-4-bromo-3-fluorobenzoic acid (6.50 g, 27.77mmol, 1.00 eq) in DMF (92.00 mL) at 25° C., was added1-chloropyrrolidine-2,5-dione (4.08 g, 30.55 mmol, 1.10 eq). The mixturewas stirred at 70° C. for 2 h. The mixture was cooled to 25° C. andpoured into cold-water (30 mL). The residue was extracted with ethylacetate (30 mL×3). The combined organic layer was washed with brine (40mL), dried over Na₂SO₄, filtered and concentrated to afford the desiredproduct (6.20 g, 23.09 mmol, 83.15% yield) as a yellow solid which wasused directly in the next step.

Methyl 2-amino-4-bromo-5-chloro-3-fluorobenzoate

To a solution of 2-amino-4-bromo-5-chloro-3-fluorobenzoic acid (3.00 g,11.17 mmol, 1.00 eq) in MeOH (50.00 mL), was added H₂SO₄ (22.08 g,225.08 mmol, 12.00 mL, 20.15 eq) slowly. The mixture was stirred at 90°C. for 16 h. The reaction mixture was concentrated under reducedpressure to remove CH₃OH, The residue was neutralized with aq.NaOH (1M)to pH=8, and then extracted with CH₂Cl₂ (15 mL×3). The combined organiclayers were washed with brine (20 mL), dried over Na₂SO₄, filtered andconcentrated in vacuo to afford the desired product (2.50 g, crude) as ayellow solid. ¹H NMR (400 MHz, DMSO-d₆): δ 7.68 (s, 1H), 6.87 (s, 2H),3.83 (s, 3H).

Methyl 3-(methylamino)propanoate

A mixture of methanamine (2 M in THF, 96.50 mL, 1.00 eq) and methylacrylate (16.80 g, 195.14 mmol, 17.50 mL, 1.01 eq) was stirred at 25° C.for 12 h. The crude compound (22.00 g, crude) in THF (96.5 mL) was useddirectly in the next step.

Methyl 3-((3-methoxy-3-oxopropyl)(methyl)amino)-3-oxopropanoate

To a solution of methyl 3-(methylamino)propanoate (22.00 g, 187.79 mmol,1.00 eq) in THF (96.50 mL), was added Et₃N (19.20 g, 189.67 mmol, 26.30mL, 1.01 eq). The mixture was cooled to 0° C. in an ice-bath, methyl3-chloro-3-oxo-propanoate (26.40 g, 193.42 mmol, 20.62 mL, 1.03 eq) wasadded drop-wise and the mixture was stirred at 0° C. for 3 h. Thereaction mixture was warmed to 25° C. and adjusted pH to 2˜3 with aq.HCl(2M). The mixture was extracted with DCM (100 mL×3). The combinedorganic layers were washed with brine (50 mL), dried over Na₂SO₄ andconcentrated under reduced pressure to give the desired product (27.00g, crude) as a yellow oil. LCMS (M+H⁺) m/z: calcd. 218.10, found 218.0.

Methyl 1-methyl-2,4-dioxopiperidine-3-carboxylate

Na (10.58 g, 460.35 mmol, 10.91 mL, 5.00 eq) was added to MeOH (120.00mL), and then methyl3-((3-methoxy-3-oxopropyl)(methyl)amino)-3-oxopropanoate (20.00 g, 92.07mmol, 1.00 eq) in Toluene (150.00 mL) was added to the above solution.The mixture was stirred at 110° C. for 2 h. The reaction mixture wascooled to 25° C. and adjusted pH to 2˜3 with aq.HCl (2M). The mixturewas extracted with EA (100 mL×3). The combined organic layers werewashed with brine (50 mL), dried over Na₂SO₄ and concentrated underreduced pressure to give the desired product which was used directly inthe next step. LCMS (M+H+) m/z: calcd. 186.07, found 185.9. ¹H NMR (400MHz, CHLOROFORM-d): δ 3.85 (s, 3H), 3.38-3.29 (m, 2H), 2.96 (s, 3H),2.66-2.61 (m, 3H).

1-Methylpiperidine-2,4-dione

Methyl 1-methyl-2,4-dioxopiperidine-3-carboxylate (15.00 g, 81.00 mmol,1.00 eq) was dissolved in H₂O (150.00 mL) and CH₃CN (150.00 mL). Themixture was heated at 100° C. for 0.5 h. The reaction mixture was cooledto 25° C. and extracted with EA (100 mL×3). The combined organic layerswere washed with brine (50 mL), dried over Na₂SO₄ and concentrated underreduced pressure to give a residue. The residue was purified by columnchromatography on silica (PE/EA=1:3˜0:1) to afford the desired product(2.50 g, 19.66 mmol, 24.28% yield) as a yellow oil. 1H NMR (400 MHz,CHLOROFORM-d): δ 3.57-3.54 (m, 2H), 3.3 (s, 2H), 3.05 (s, 3H), 2.65-2.62(m, 2H).

7-Bromo-8-chloro-6-fluoro-10-hydroxy-2-methyl-3,4-dihydrobenzo[b][1,6]naphthyridin-1(2H)-one

To a mixture of 1-methylpiperidine-2,4-dione (2.01 g, 15.77 mmol, 1.00eq) and methyl 2-amino-4-bromo-5-chloro-3-fluorobenzoate (4.50 g, 15.93mmol, 1.01 eq) in toluene (50.00 mL), was added CH₃SO₃H (9.45 g, 98.26mmol, 7.00 mL, 6.23 eq). The mixture was stirred at 95° C. for 16 h. Themixture was cooled to 25° C. and concentrated under reduced pressure toremove toluene. The residue was adjusted pH to 7˜8 with aq.NaOH (2M) togive the desired product (2.00 g, 5.56 mmol, 35.26% yield) as a yellowsolid. LCMS (M+H⁺) m/z: calcd. 358.95, found 358.9. ¹H NMR (400 MHz,DMSO-d₆): δ 8.08 (s, 1H), 3.66-3.65 (m, 2H), 3.14-3.11 (m, 2H), 3.03 (s,3H).

7-Bromo-8,10-dichloro-6-fluoro-2-methyl-3,4-dihydrobenzo[b][1,6]naphthyridin-1(2H)-one

A mixture of7-bromo-8-chloro-6-fluoro-10-hydroxy-2-methyl-3,4-dihydrobenzo[b][1,6]naphthyridin-1(2H)-one(850.00 mg, 2.36 mmol, 1.00 eq) in POCl3 (16.50 g, 107.61 mmol, 10.00mL, 45.60 eq) was stirred at 120° C. for 6 h. The reaction solution wasconcentrated under reduced pressure to give a residue. Then the residuewas cooled to 25° C., adjusted pH to 7˜8 with saturated NaHCO₃ solutionand extracted with EA (30 mL×3). The combined organic layers were washedwith brine (20 mL), dried over Na₂SO₄, filtered and concentrated underreduced pressure to give the desired product (700.00 mg, crude) as a redbrown solid. LCMS (M+H⁺) m/z: calcd. 376.92, found 376.9. ¹H NMR (400MHz, CHLOROFORM-d): δ 8.40 (s, 1H), 3.72-3.70 (m, 2H), 3.42-3.39 (m,2H), 3.26 (s, 3H).

tert-Butyl4-(7-bromo-8-chloro-6-fluoro-2-methyl-1-oxo-1,2,3,4-tetrahydrobenzo[b][1,6]naphthyridin-10-yl)piperazine-1-carboxylate

To a mixture of7-bromo-8,10-dichloro-6-fluoro-2-methyl-3,4-dihydrobenzo[b][1,6]naphthyridin-1(2H)-one(700.00 mg, 1.85 mmol, 1.00 eq) and tert-butyl piperazine-1-carboxylate(410.00 mg, 2.20 mmol, 1.19 eq) in toluene (40.00 mL), was added Et₃N(730.00 mg, 7.21 mmol, 1.00 mL, 3.90 eq). The mixture was stirred at 25°C. for 3 h. The reaction mixture was concentrated under reduced pressureto remove toluene. The residue was diluted with H₂O (20 mL) andextracted with ethyl acetate (30 mL×3). The combined organic layers waswashed with brine (20 mL), dried over Na₂SO₄, filtered and concentratedunder reduced pressure to give a residue. The residue was purified byflash silica gel chromatography (PE/EA=1:1˜1.5:1) to afford the desiredproduct (300.00 mg, 528.60 umol, 28.57% yield) as a red solid. LCMS(M+H⁺⁾m/z: calcd. 527.08, found 527.1. ¹H NMR (400 MHz, CHLOROFORM-d): δ8.09 (s, 1H), 3.69-3.65 (m, 4H), 3.64-3.62 (m, 2H), 3.35-3.34 (m, 4H),3.29-3.26 (m, 2H), 3.24 (s, 3H), 1.50 (s, 9H).

tert-Butyl4-(8-chloro-6-fluoro-7-(2-fluoro-6-hydroxyphenyl)-2-methyl-1-oxo-1,2,3,4-tetrahydrobenzo[b][1,6]naphthyridin-10-yl)piperazine-1-carboxylate

To a mixture of tert-Butyl4-(7-bromo-8-chloro-6-fluoro-2-methyl-1-oxo-1,2,3,4-tetrahydrobenzo[b][1,6]naphthyridin-10-yl)piperazine-1-carboxylate(100.00 mg, 189.46 umol, 1.00 eq), (2-fluoro-6-hydroxyphenyl)boronicacid (35.00 mg, 224.47 umol, 1.18 eq), RuPhos (8.84 mg, 18.95 umol, 0.10eq) and K₂CO₃ (52.37 mg, 378.92 umol, 2.00 eq) in the co-solvent ofdioxane (8.00 mL) and H₂O (2.00 mL), was added Ruphos Pd G₃ (8.00 mg,9.57 umol, 0.05 eq). The resulting mixture was degassed and purged withN₂ for 3 cycles. The reaction was heated at 85° C. under N₂ atmospherefor 7 h. The reaction mixture was concentrated under reduced pressure togive a residue. The residue was purified by prep-TLC (SiO2, PE/EA=1:1.5)to afford the desired product (25.00 mg, 40.25 umol, 10.62% yield) as alight yellow solid. LCMS (M+H⁺⁾m/z: calcd. 559.18, found 559.0. ¹H NMR(400 MHz, CHLOROFORM-d): δ 8.00 (s, 1H), 7.35-7.29 (m, 1H), 6.87-6.85(m, 1H), 6.79-6.75 (m, 1H), 3.71-3.66 (m, 4H), 3.63-3.62 (m, 2H),3.62-3.59 (m, 2H), 3.30-3.26 (m, 4H), 3.23 (s, 3H), 1.50 (s, 9H).

8-Chloro-6-fluoro-7-(2-fluoro-6-hydroxyphenyl)-2-methyl-10-(piperazin-1-yl)-3,4-dihydrobenzo[b][1,6]naphthyridin-1(2H)-one

To a mixture of tert-butyl4-(8-chloro-6-fluoro-7-(2-fluoro-6-hydroxyphenyl)-2-methyl-1-oxo-1,2,3,4-tetrahydrobenzo[b][1,6]naphthyridin-10-yl)piperazine-1-carboxylate(20.00 mg, 35.78 umol, 1.00 eq) in DCM (10.00 mL), was added CF₃COOH(1.54 g, 13.51 mmol, 1.00 mL, 377.48 eq). The resulting mixture wasstirred at 0° C. for 5 h. The reaction mixture was concentrated underreduced pressure to give the desired product (15.00 mg, crude) as ayellow solid which was used directly in the next step.

10-(4-acryloylpiperazin-1-yl)-8-chloro-6-fluoro-7-(2-fluoro-6-hydroxyphenyl)-2-methyl-3,4-dihydrobenzo[b][1,6]naphthyridin-1(2H)-one

To a solution of8-Chloro-6-fluoro-7-(2-fluoro-6-hydroxyphenyl)-2-methyl-10-(piperazin-1-yl)-3,4-dihydrobenzo[b][1,6]naphthyridin-1(2H)-one (15.00 mg, 32.69 umol, 1.00 eq) and Et₃N (33.08 mg, 326.90umol, 45.32 uL, 10.00 eq) in DCM (5.00 mL), was added prop-2-enoylchloride (1.48 mg, 16.35 umol, 1.33 uL, 0.50 eq) at 0° C. The resultingmixture was stirred at 0° C. for 15 min. The reaction mixture wasquenched with saturated NaHCO₃ (5 mL). The mixture was extracted withDCM (3×5 mL). The organic layer was washed with brine (5 mL), dried withNa₂SO₄ and concentrated under reduced pressure to give a residue. Theresidue was purified by prep-TLC (SiO2, DCM/MeOH=20:1) to afford thedesired product (7.9 mg, 12.17 umol, 37.21% yield) as a yellow solid.LCMS (M+H⁺⁾ m/z: calcd. 513.14, found 513.1. ¹H NMR (400 MHz,CHLOROFORM-d): δ 8.00 (s, 1H), 7.32-7.28 (m, 1H), 6.88-6.86 (m, 1H),6.75-6.71 (m, 1H), 6.61-6.57 (m, 1H), 6.35-6.31 (m, 1H), 5.76-5.73 (m,1H), 3.90-3.77 (m, 4H), 3.62-3.57 (m, 2H), 3.30-3.24 (m, 2H), 3.21-3.10(m, 4H), 3.08 (s, 3H).

Example 2 Synthesis of5-(4-acryloylpiperazin-1-yl)-7-chloro-9-fluoro-8-(2-fluoro-6-hydroxyphenyl)-3,4-dihydropyrimido[4,5-b]quinolin-2(1h)-one

Example 2 provides an exemplary preparation according to Method B.

7-Bromo-6-chloro-8-fluoro-1H-3,1-benzoxazine-2,4-dione

To a suspension of compound 2-amino-4-bromo-5-chloro-3-fluorobenzoicacid (500 mg, 1.86 mmol, 1.00 eq) in THF (6.00 mL), was addedTRIPHOSGENE (187.66 mg, 632.40 umol, 0.34 eq) slowly. The mixture wasstirred at 25° C. for 4 h. The mixture was added into PE (5 mL). Theprecipitate was collected by filtration and dried to afford the desiredproduct (500 mg, 1.7 mmol, crude product) as a yellow solid. ¹H NMR (400MHz, CDCl₃): δ 8.03 (s, 1H). The yellow solid was used in the next stepdirectly.

2-Amino-7-bromo-6-chloro-8-fluoro-4-hydroxy-quinoline-3-carbonitrile

To the solution of compound7-bromo-6-chloro-8-fluoro-2H-benzo[d][1,3]oxazine-2,4(1H)-dione (830.00mg, 2.82 mmol, 1.00 eq) in DMF (12.00 mL), were added propanedinitrile(372.41 mg, 5.64 mmol, 354.68 uL, 2.00 eq) and DIPEA (728.58 mg, 5.64mmol, 984.57 uL, 2.00 eq) sequentially. The resulting solution wasstirred at 95° C. for 1 h. Most of solvent was removed in vacuo. Theresidue was treated with water (20 mL) and solid precipitated out. Thesolid was filtered and washed with water and PE to give the desiredproduct (1.00 g, crude) as a grey solid. ¹H NMR (400 MHz, DMSO-d₆): δ7.33 (s, 1H), 5.85 (s, 2H).

2-Amino-7-bromo-4,6-dichloro-8-fluoro-quinoline-3-carbonitrile

A solution of compound2-amino-7-bromo-6-chloro-8-fluoro-4-hydroxy-quinoline-3-carbonitrile(1.00 g, 3.16 mmol, 1.00 eq) in POCl₃ (19.80 g, 129.13 mmol, 12.00 mL,40.86 eq) was stirred at 100° C. under N₂ atmosphere for 16.5 h. Thesolution turned to a yellow suspension. The mixture was poured intowater (80 mL) and stirred for 5 min. The mixture was adjusted to pH=8˜9with 4M NaOH. The mixture was treated with EA (80 mL) and filtered toremove some unsoluable brown solid. The filtrate was separated and theaqueous was extracted with EA (80 mL×2). The combined organic layerswere washed with brine, dried over anhydrous Na₂SO₄, filtered andconcentrated to give the desired product (979.00 mg, crude) as a redsolid. The crude product was used in the next step directly.

tert-Butyl4-(2-amino-7-bromo-6-chloro-3-cyano-8-fluoro-4-quinolyl)piperazine-1-carboxylate

To the solution of compound2-amino-7-bromo-4,6-dichloro-8-fluoro-quinoline-3-carbonitrile (979.00mg, 2.92 mmol, 1.00 eq) and tert-butyl piperazine-1-carboxylate (816.54mg, 4.38 mmol, 1.50 eq) in toluene (15.00 mL), was added TEA (591.50 mg,5.84 mmol, 810.27 uL, 2.00 eq). The resulting mixture was stirred at110° C. for 9.5 h. The reaction mixture was cooled down and diluted withEA (50 mL). The mixture was washed with water (30 mL) and separated. Theaqueous phase was extracted with EA (20 mL×2). The combined organiclayers were washed with brine, dried over anhydrous Na₂SO₄, filtered andconcentrated. The residue was purified by column chromatography onsilica gel (PE/EA=3/1˜2/1) to give the desired product (428.00 mg,882.93 umol, 30.24% yield) as a yellow solid. LCMS (M+H⁺) m/z: calcd.484.05, found 484.0, 486.0. ¹H NMR (400 MHz, CDCl₃): δ 7.69 (s, 1H),3.72 (s, 4H), 3.59 (t, 4H), 1.52 (s, 9H).

tert-Butyl4-[2-amino-6-chloro-3-cyano-8-fluoro-7-(2-fluoro-6-methoxy-phenyl)-4-quinolyl]piperazine-1-carboxylate

To the mixture of tert-Butyl4-(2-amino-7-bromo-6-chloro-3-cyano-8-fluoro-4-quinolyl)piperazine-1-carboxylate(300.00 mg, 618.88 umol, 1.00 eq), (2-fluoro-6-methoxy-phenyl)boronicacid (126.21 mg, 742.66 umol, 1.20 eq), K₂CO₃ (5.34 mg, 38.62 umol, 2.00eq) and RuPhos (901.23 ug, 1.93 umol, 0.10 eq) in the co-solvent ofdioxane (9.00 mL) and Water (2.25 mL), was added RuPhos Pd G3 (1.62 mg,1.93 umol, 0.10 eq). The resulting mixture was degassed and purged withN₂ for 3 cycles. The reaction was heated at 80° C. under N₂ atmospherefor 30 min. The reaction was diluted with EA (10 mL) and washed withbrine (8 mL). The organic layer was dried over anhydrous Na₂SO₄,filtered and concentrated. The residue was purified by columnchromatography on silica gel (PE/EA=4/1˜2/1) to give the desired product(256.00 mg, 396.10 umol, 64.00% yield) as a yellow solid. LCMS (M+H⁺)m/z: calcd. 530.17, found 530.1. ¹H NMR (400 MHz, CDCl₃): δ 7.72 (d,1H), 7.46-7.40 (m, 1H), 6.88-6.83 (m, 2H), 5.65 (s, 1H), 3.80 (s, 3H),3.73 (s, 4H), 3.64-3.62 (m, 4H), 1.51 (s, 9H).

tert-Butyl4-[2-amino-3-(aminomethyl)-6-chloro-8-fluoro-7-(2-fluoro-6-methoxy-phenyl)-4-quinolyl]piperazine-1-carboxylate

To a solution of tert-butyl4-[2-amino-6-chloro-3-cyano-8-fluoro-7-(2-fluoro-6-methoxy-phenyl)-4-quinolyl]piperazine-1-carboxylate(150.00 mg, 283.03 umol, 1.00 eq) in MeOH (6.00 mL) and THF (2.00 mL),was added NiCl₂.6H₂O (67.27 mg, 283.03 umol, 1.00 eq). NaBH₄ (214.14 mg,5.66 mmol, 20.00 eq) was added into the mixture over 15 min. The mixturewas stirred at 25° C. for 30 min. The mixture was concentrated. Theresidue was diluted with water (15 mL), extracted with DCM (3×30 mL).The combined organic layer was washed with brine (30 mL), dried overNa₂SO₄, filtered and concentrated. The residue was purified by prep-TLC(DCM/MeOH=20/1) to afford the desired product (100.00 mg, 187.27 umol,66.16% yield) as a yellow solid.

tert-Butyl4-[7-chloro-9-fluoro-8-(2-fluoro-6-methoxy-phenyl)-2-oxo-3,4-dihydro-1H-pyrimido[4,5-b]quinolin-5-yl]piperazine-1-carboxylate

To a solution of tert-butyl4-[2-amino-3-(aminomethyl)-6-chloro-8-fluoro-7-(2-fluoro-6-methoxy-phenyl)-4-quinolyl]piperazine-1-carboxylate(100.00 mg, 187.27 umol, 1.00 eq) in DMF (3.00 mL), was added to CDI(33.40 mg, 206.00 umol, 1.10 eq). The mixture was stirred at 25° C. for16 h. The mixture was concentrated in vacuo. The mixture was purified byprep-TLC (DCMICH₃OH=20:1) to afford the desired product (100.00 mg,169.65 umol, 90.59% yield) as a yellow solid. LCMS (M+H⁺) m/z: calcd.560.18, found 560.2.

7-Chloro-9-fluoro-8-(2-fluoro-6-methoxy-phenyl)-5-piperazin-1-yl-3,4-dihydro-1H-pyrimido[4,5-b]quinolin-2-one

To a solution of tert-Butyl4-[7-chloro-9-fluoro-8-(2-fluoro-6-methoxy-phenyl)-2-oxo-3,4-dihydro-1H-pyrimido[4,5-b]quinolin-5-yl]piperazine-1-carboxylate(100.00 mg, 178.57 umol, 1.00 eq) in DCM (5.00 mL), was added CF₃COOH(770.00 mg, 6.75 mmol, 500.00 uL, 37.82 eq) at 0° C. The mixture wasstirred at 25° C. for 3 h. The mixture was concentrated in vacuo.Compound 14 (80 mg, crude) was obtained as a yellow oil. The crudeproduct was used directly in the next step. LCMS (M+H⁺) m/z: calcd.459.9, found 460.1, 461.1.

7-Chloro-9-fluoro-8-(2-fluoro-6-methoxy-phenyl)-5-(4-prop-2-enoylpiperazin-1-yl)-3,4-dihydro-1H-pyrimido[4,5-b]quinolin-2-one

To a solution of7-Chloro-9-fluoro-8-(2-fluoro-6-methoxy-phenyl)-5-piperazin-1-yl-3,4-dihydro-1H-pyrimido[4,5-b]quinolin-2-one(80.00 mg, 173.96 umol, 1.00 eq) in DCM (5.00 mL), was added Et₃N (52.81mg, 521.88 umol, 72.34 uL, 3.00 eq) at 0° C. ACRYLOYL CHLORIDE (15.74mg, 173.96 umol, 14.18 uL, 1.00 eq) was added to the mixture slowly at0° C. The mixture was stirred at 0° C. for 0.5 h. Water (3 mL) was addedto the mixture. The mixture was extracted with DCM (3 mL*3). Thecombined organic layers were dried over Na₂SO₄, filtered andconcentrated in vacuo. The mixture was purified by prep-TLC(DCM/CH3OH=20:1) to afford the desired product (70.00 mg, 133.48 umol,76.73% yield) as a white solid. LCMS (M+H⁺) m/z: calcd. 514.14, found514.1.

7-Chloro-9-fluoro-8-(2-fluoro-6-hydroxy-phenyl)-5-(4-prop-2-enoylpiperazin-1-yl)-3,4-dihydro-1H-pyrimido[4,5-b]quinolin-2-one

To a solution of7-Chloro-9-fluoro-8-(2-fluoro-6-methoxy-phenyl)-5-(4-prop-2-enoylpiperazin-1-yl)-3,4-dihydro-1H-pyrimido[4,5-b]quinolin-2-one(35.00 mg, 68.10 umol, 1.00 eq) in DCM (5.00 mL), was added BBr₃ (341.21mg, 1.36 mmol, 131.23 uL, 20.00 eq) at −78° C. under N₂ atmosphere. Themixture was stirred at 25° C. for 2 h. Saturated aq. NaHCO₃ (3 mL) wasadded to the mixture at −78° C. The mixture was concentrated in vacuo.The residue was extracted with EA (3 mL×3). The combined organic layerswere dried over Na₂SO₄, filtered and concentrated in vacuo. The mixturewas purified by prep-TLC (DCM/CH₃OH=15:1) to afford the desired product(4.90 mg, 8.53 umol, 12.52% yield) as a light yellow solid. LCMS (M+H⁺)m/z: calcd. 500.12, found 499.9. ¹H NMR (400 MHz, DMSO-d₆): δ 10.19 (s,1H), 10.05 (s, 1H), 7.96 (s, 1H), 7.38-7.28 (m, 1H), 7.28-7.19 (m, 1H),6.94-6.87 (m, 1H), 6.84 (d, J=8.8 Hz, 1H), 6.79 (t, J=8.8 Hz, 1H), 6.17(dd, J=16.5, 2.0 Hz, 1H), 5.73 (dd, J=10.4, 2.0 Hz, 1H), 4.57 (s, 2H),4.07-3.48 (m, 4H), 3.29-3.09 (m, 4H).

Example 3 Biochemical Assay of the Compounds

Test compounds are prepared as 10 mM stock solutions in DMSO (Fisher cat# BP-231-100). KRAS G12C 1-169, his-tagged protein, GDP-loaded isdiluted to 2 μm in buffer (20 mM Hepes, 150 mM NaCl, 1 mM MgCl₂).Compounds are tested for activity as follows:

Compounds are diluted to 50× final test concentration in DMSO in 96-wellstorage plates. Compound stock solutions are vortexed before use andobserved carefully for any sign of precipitation. Dilutions are asfollows:

-   -   For 100 μM final compound concentration, compounds are diluted        to 5000 μM (5 μl 10 mM compound stock+5 μl DMSO and mixed well        by pipetting.    -   For 30 μM final compound concentration, compounds are diluted to        1500 μM (3 μl 10 mM compound stock+17 μl DMSO) and mixed well by        pipetting.    -   For 10 μM final compound concentration, compounds are diluted to        500 μM (2 μl 10 mM compound stock+38 μl DMSO) and mixed well by        pipetting.        49 μl of the stock protein solution is added to each well of a        96-well PCR plate (Fisher cat #1423027). 141 of the diluted 50×        compounds is added to appropriate wells in the PCR plate using        12-channel pipettor. Reactions are mixed carefully and        thoroughly by pipetting up/down with a 200 μl multi-channel        pipettor. The plate is sealed well with aluminum plate seal, and        stored in a drawer at room temperature for 30 min, 2 hour or 24        hrs. 5 μl of 2% formic acid (Fisher cat # A117) in DI H₂O is        then added to each well followed by mixing with a pipette. The        plate is then resealed with aluminum seal and stored on dry ice        until analyzed as described below.

The above described assays are analyzed by mass spectrometry accordingto one of the following two procedures:

RapidFire/TOF Assay:

The MS instrument is set to positive polarity, 2 GHz resolution, and lowmass (1700) mode and allowed to equilibrate for 30 minutes. Theinstrument is then calibrated, switched to acquisition mode and theappropriate method loaded.

After another 30 minute equilibration time, a blank batch (i.e., buffer)is run to ensure equipment is operating properly. The samples are thawedat 37° C. for 10 minutes, briefly centrifuged, and transferred to thebench top. Wells A1 and H12 are spiked with 1 uL 500 uM internalstandard peptide, and the plates centrifuged at 2000×g for 5 minutes.The method is then run and masses of each individual well recorded.

The masses (for which integration data is desired) for each well arepasted into the platemap and exported from the analysis. Masses for theinternal standards are exported as well. The data at 50 ppm is extractedfor the +19 charge state, and identity of well A1 is assigned using theinternal standard spike and integrated. Peak data is exported as a TOFlist and the above steps are repeated individually, for the +20, 21, 22,23, 24, and 25 charge states.

Q-Exactive Assay:

The masses and peak intensities of KRAS G12C protein species aremeasured using a Dionex RSLCnano system (Thermo Scientific) connected toa Q Exactive Plus mass spectrometer (Thermo Scientific).

20 mL of sample are each loaded onto a Aeris™ 3.6 m WIDEPORE C4 200 Å,LC Column 50×2.1 mm column maintained at 40° C. at a flow rate of 600 μlmin⁻¹ with 20% Solvent A (0.1% formic acid in H₂O) and 80% Solvent B(0.1% formic acid in acetonitrile). The liquid chromatography conditionsare 20% solvent B for 1 min, 20% to 60% solvent B for 1.5 min, 60% to90% solvent for 0.5 min, 90% solvent B for 0.2 min, 90% to 20% solvent Bfor 0.2 min, and then equilibrated for 1.6 min before the followingsample injection. The flow rate is maintained at 600 μl min⁻¹ throughoutthe sample analysis.

The mass spectrometer is operated in profile mode at a resolution of17500, 5 microscans, using 50 msec max injection time and an AGC targetof 1e6, and a full mass range from 800-1850 m/z is recorded. The HCDtrapping gas is optimized for maximum sensitivity for intact proteins.The ionization method is electrospray ionization, which uses a sprayvoltage of 4 kV, sheath gas flow set to 50 au, auxiliary gas flow set to10 au and sweep gas flow set to 1 au. The capillary ion transfertemperature was 320° C. and the S-lens RF level was set to 50 voltage.Protein Deconvolution software (Thermo Scientific) was used todeconvolute the charge envelopes of protein species in samples.

Data is analyzed using the Thermo protein deconvolution package.Briefly, the charge envelope for each observed species is quantitativelydeconvoluted to determine the mass and intensity of each parent species(modified or unmodified protein). % modification is calculated based onthe deconvoluted peak intensities.

Other in vitro analyses are as follows:

Inhibition of Cell Growth:

The ability of the subject compounds to inhibit RAS-mediated cell growthis assessed and demonstrated as follows. Cells expressing a wildtype ora mutant RAS are plated in white, clear bottom 96 well plates at adensity of 5,000 cells per well. Cells are allowed to attach for about 2hours after plating before a compound disclosed herein is added. Aftercertain hours (e.g., 24 hours, 48 hours, or 72 hours of cell growth),cell proliferation is determined by measuring total ATP content usingthe Cell Titer Glo reagent (Promega) according to manufacturer'sinstructions. Proliferation EC50s is determined by analyzing 8 pointcompound dose responses at half-log intervals decreasing from 100 μM.

Inhibition of RAS-Mediated Signaling Transduction:

The ability of the compounds disclosed herein in inhibiting RAS-mediatedsignaling is assessed and demonstrated as follows. Cells expressing wildtype or a mutant RAS (such as G12C, G12V, or G12A) are treated with orwithout (control cells) a subject compound. Inhibition of RAS signalingby one or more subject compounds is demonstrated by a decrease in thesteady-state level of phosphorylated MEK, phosphorylated ERK,phosphorylated RSK, and/or Raf binding in cells treated with the one ormore of the subject compounds as compared to the control cells.

Exemplary compounds of structure (I), including exemplary compounds fromTable 1 are tested according to the above methods and found tocovalently bind to KRAS G12C to the extent of at least about 5% after 2hour incubation (i.e., at least about 5% of the protein present in thewell was found to be covalently bound to test compound). Results forexemplary compounds are provided in Table 2.

TABLE 2 Activity of Representative Compounds of Structure (I) No.Binding % No. Binding % 1 ++ 19 + + indicates binding activity up to to50% ++ indicates binding activity greater than 50%

All of the U.S. patents, U.S. patent application publications, U.S.patent applications, foreign patents, foreign patent applications andnon-patent publications referred to in this specification or theattached Application Data Sheet are incorporated herein by reference, intheir entirety to the extent not inconsistent with the presentdescription.

U.S. provisional patent application Ser. No. 62/147,955 filed Apr. 15,2015 entitled “FUSED-TRICYCLIC INHIBITORS OF KRAS AND METHODS OF USETHEREOF” is incorporated herein by reference, in its entirety.

From the foregoing it will be appreciated that, although specificembodiments of the invention have been described herein for purposes ofillustration, various modifications may be made without deviating fromthe spirit and scope of the invention. Accordingly, the invention is notlimited except as by the appended claims.

The invention claimed is:
 1. A compound having the following structure(I′a):

or a pharmaceutically acceptable salt, stereoisomer or prodrug thereof,wherein: A is a carbocyclic, heterocyclic or heteroaryl ring; G¹ and G²are each independently N or CH; L¹ is a bond or NR⁵; L² is a bond oralkylene; R¹ is aryl or heteroaryl; R^(2a), R^(2b) and R^(2c) are eachindependently H, amino, halo, hydroxyl, cyano, C₁-C₆ alkyl, C₁-C₆alkylaminyl, C₁-C₆ haloalkyl, C₁-C₆ alkoxy, C₁-C₆ haloalkoxy; C₃-C₈cycloalkyl, heterocyclylalkyl, C₂-C₆ alkynyl, C₂-C₆ alkenyl,aminylalkyl, alkylaminylalkyl, cyanoalkyl, carboxyalkyl,aminylcarbonylalkyl, aminylcarbonyl, heteroaryl or aryl; R^(3a) andR^(3b) are, at each occurrence, independently H, —OH, —NH₂, —CO₂H, halo,cyano, C₁-C₆ alkyl, C₁-C₆ haloalkyl, C₁-C₆ haloalkoxy, C₃-C₈ cycloalkyl,heterocyclylalkyl, C₂-C₆ alkynyl, hydroxylalkyl, alkoxyalkyl,aminylalkyl, alkylaminylalkyl, cyanoalkyl, carboxyalkyl,aminylcarbonylalkyl or aminylcarbonyl; or R^(3a) and R^(3b) join to forma carbocyclic or heterocyclic ring; or R^(3a) is H, —OH, —NH₂, —CO₂H,halo, cyano, C₁-C₆ alkyl, C₁-C₆ haloalkyl, C₁-C₆ haloalkoxy, C₃-C₈cycloalkyl, heterocyclylalkyl, C₂-C₆ alkynyl, hydroxylalkyl,alkoxyalkyl, aminylalkyl, alkylaminylalkyl, cyanoalkyl, carboxyalkyl,aminylcarbonylalkyl or aminylcarbonyl, and R^(3b) joins with R^(4b) toform a carbocyclic or heterocyclic ring; R^(4a) and R^(4b) are, at eachoccurrence, independently H, —OH, —NH₂, —CO₂H, halo, cyano, C₁-C₆ alkyl,C₁-C₆ haloalkyl, C₁-C₆ haloalkoxy, C₃-C₈ cycloalkyl, heterocyclylalkyl,C₂-C₆ alkynyl, hydroxylalkyl, alkoxyalkyl, aminylalkyl,alkylaminylalkyl, cyanoalkyl, carboxyalkyl, aminylcarbonylalkyl oraminylcarbonyl; or R^(4a) and R^(4b) join to form a carbocyclic orheterocyclic ring; or R^(4a) is H, —OH, —NH₂, —CO₂H, halo, cyano, C₁-C₆alkyl, C₁-C₆ haloalkyl, C₁-C₆ haloalkoxy, C₃-C₈ cycloalkyl,heterocyclylalkyl, C₂-C₆ alkynyl, hydroxylalkyl, alkoxyalkyl,aminylalkyl, alkylaminylalkyl, cyanoalkyl, carboxyalkyl,aminylcarbonylalkyl or aminylcarbonyl, and R^(4b) joins with R^(3b) toform a carbocyclic or heterocyclic ring; R⁵ is, at each occurrence,independently H, C₁-C₆ alkyl, C₃-C₈ cycloalkyl or heterocycloalkyl; m¹and m² are each independently 1, 2 or 3;

represents a double or triple bond; Q is —C(═O)—, —C(═NR^(8′))—,NR⁸C(═O)—, —S(═O)₂— or NR⁸S(═O)₂; R⁸ is H, C₁-C₆ alkyl, hydroxylalkylaminoalkyl, alkoxyalkyl, aminylalkyl, alkylaminylalkyl, cyanoalkyl,carboxyalkyl, aminylcarbonylalkyl, C₃-C₈ cycloalkyl or heterocycloalkyl;R^(8′) is H, —OH, —CN or C₁-C₆ alkyl; when

is a double bond then R⁹ and R¹⁰ are each independently H, halo, cyano,carboxyl, C₁-C₆ alkyl, alkoxycarbonyl, aminylalkyl, alkylaminylalkyl,aryl, heterocyclyl, heterocyclylalkyl, heteroaryl or hydroxylalkyl, orR⁹ and R¹⁰ join to form a carbocyclic, heterocyclic or heteroaryl ring;and when

is a triple bond then R⁹ is absent and R¹⁰ is H, C₁-C₆, alkyl,aminylalkyl, alkylaminylalkyl or hydroxylalkyl.
 2. The compound of claim1, wherein the compound has one of the following structures (I′b),(I′c), (I'd) or (I′e):


3. The compound of claim 1, wherein A is a 5, 6 or 7-memberedheterocyclic or heteroaryl ring.
 4. The compound of claim 1, wherein Ahas one of the following structures:

wherein: R⁶ is, at each occurrence, independently H, halo, amino, cyano,arayl, heteroaryl, C₁-C₆ alkyl, C₃-C₅ cycloalkyl, heterocycloalkyl,hydroxylalkyl, alkoxyalkyl, aminylalkyl, alkylaminylalkyl, cyanoalkyl,carboxyalkyl, aminylcarbonylalkyl, arylalkyl or heteroarylalkyl; X is Oor CH₂; and n is 0, 1 or
 2. 5. The compound of claim 1, wherein R¹ isaryl.
 6. The compound of claim 5, wherein R¹ is phenyl or naphthyl. 7.The compound of claim 5, wherein R¹ is substituted with halo, aminohydroxyl, C₁-C₆ alkyl, cyano, C₁-C₆ haloalkyl, C₁-C₆ alkoxy,alkylaminyl, cycloalkyl, heterocyclylalkyl, aryl, heteroaryl, boronicacid, —OC(═O)R, phosphate, phosphoalkoxy or C₁-C₆ alkylcarbonyloxy, orcombinations thereof, wherein R is C₁-C₆ alkyl.
 8. The compound of claim7, wherein R¹ is substituted with fluoro, chloro, hydroxyl, methyl,isopropyl, cyclopropyl, trifluoromethyl or methoxy, or combinationsthereof.
 9. The compound of claim 1, wherein R¹ is heteroaryl.
 10. Thecompound of claim 9, wherein R¹ is indazolyl, indolyl, benzimidazolyl,benzotriazolyl or quinolinyl.
 11. The compound of claim 9, wherein R¹ issubstituted with hydroxyl, halo or C₁-C₆ alkyl, or combinations thereof.12. The compound of claim 1, wherein R¹ has one of the followingstructures:


13. The compound of claim 1, wherein R^(2c) is H.
 14. The compound ofclaim 1, wherein R^(2a) and R^(2b) are each independently halo,haloalkyl, alkyl or alkoxy.
 15. The compound of claim 1, wherein Q isC(═O)—.
 16. The compound of claim 1, wherein R⁹ and R¹⁰ are each H. 17.The compound of claim 1, wherein the moiety

has one of the following structures:


18. The compound of claim 1, wherein the moiety has the followingstructure:


19. The compound of claim 1, wherein L¹ is a bond.
 20. The compound ofclaim 1, wherein L² is a bond.
 21. The compound of claim 1, whereinR^(3a), R^(3b), R^(4a) and R^(4b) are each H.
 22. The compound of claim1, wherein the compound has one of the following structures:


23. A substantially purified atropisomer of a compound according toclaim
 1. 24. A pharmaceutical composition comprising a compound of claim1 and a pharmaceutically acceptable carrier.
 25. A method for treatmentof cancer, the method comprising administering an effective amount ofthe pharmaceutical composition of claim 24 to a subject in need thereof.26. A method for regulating activity of a KRAS, HRAS or NRAS G12C mutantprotein, the method comprising reacting the KRAS G12C mutant proteinwith the compound of claim
 1. 27. A method for inhibiting proliferationof a cell population, the method comprising contacting the cellpopulation with the compound of claim
 1. 28. A method for preparing alabeled KRAS, HRAS or NRAS G12C mutant protein, the method comprisingreacting the KRAS, HRAS or NRAS G12C mutant with a compound of claim 1,to result in the labeled KRAS, HRAS or NRAS G12C protein.